ALBERT

Notes: Summary Rats were injected with45Ca and horseradish peroxidase to determine the patterns of45Ca incorporation into incisor enamel and the morphological types of the overlying maturation ameloblasts.45Ca autoradiography showed no differences in the patterns of incorporation into enamel between routinely embedded and freeze-dried specimens. Enamel overlaid by ruffle-ended ameloblasts was much more heavily labeled while that overlaid by smooth-ended ameloblasts showed only moderate labeling. The observations lend further support to the hypothesis that the ruffle-ended cells are very active in mineralizing enamel and that the smooth-ended cells are in a passive, restorative phase.

Notes: Summary Preliminary indications of the occurrence of actin and myosin in crude matrix vesicle preparations have been reported previously. In the present study extracellular matrix vesicles from rat alveolar bone were isolated. They were further purified by a sucrose density gradient. SDS-polyacrylamide gel electrophoresis of the purified vesicles revealed the presence of a polypeptide with a molecular weight of 43 K daltons and with electrophoretic mobility identical to that of blood platelet actin. The limited proteolysis of both 43 K dalton vesicular polypeptide and actin byStaphylococcus aureus-V8-protease revealed three fragments with identical electrophoretic mobility. In addition, the vesicular preparations inhibited the activity of DNase I, a property typical of actin monomers. Filamentous material extracted from matrix vesicles showed ultrastructural features of F-actin. Reaction of this material with heavy meromyosin resulted in arrowhead formation, which is characteristic of acto-heavy meromyosin. The occurrence of actin in extracellular matrix vesicles may account for their budding from the osteoblastic plasma membrane, their possible motility in the matrix, and maintenance of the spherical shape.

Notes: Summary Plasma concentrations of orphenadrine were measured by a specific gaschromatographic method in 5 healthy male volunteers after a single oral dose of orphenadrine hydrochloride 100mg. The single dose pharmacokinetic profile of orphenadrine was evaluated from these data. The elimination half-life ranged from 13.2–20.1 h after the commercial tablet formulation. Plasma concentrations, determined in volunteers and patients under different conditions of repeated oral administration of the same formulation of orphenadrine hydrochloride exceeded the theoretical values, predicted from the single dose pharmacokinetics, by a factor 2 to 3. The elimination half-lives after discontinuation of treatment showed a 2 to 3-fold increase over the single dose values. This demonstrates a clear discrepancy between the multiple and single dose pharmacokinetics of orphenadrine. Experiments in dogs suggested competition for biotransformation between orphenadrine and its metabolite N-demethylorphenadrine. Product inhibition of this type could explain the observed discrepancy.

Notes: Summary Two formulations of indapamide tablets (2.5 mg) were given as a 5.0 mg dose and the subsequent blood levels were compared to those obtained after administration of a 5.0 mg solution. The study was conducted as a randomized three-way crossover design using healthy male volunteers. The drug was well tolerated by all the subjects involved. The area under the blood concentration versus time curve, extrapolated to infinity was essentially the same for all three formulations (4.2, 4.7, and 4.4 µg-h/ml). Statistical comparison of the blood levels from the two tablets showed that one tablet had a significantly greater maximum blood concentration (263 vs 231 ng/ml) and a significantly shorter time of maximum blood concentration (2.3 vs 3.5 h). Cmax (333 ng/ml) and tmax (0.7 h) values for the solution were significantly higher than either tablet. The average half-life (β-phase) for all three formulations was 15 h, while the average systemic clearance was 20 ml/min. Indapamide has a low clearance rate and there was no evidence that the drug undergoes a first-pass effect.

Notes: Summary 21 healthy women undergoing gynaecological operations received rectal premedication with morphine 0.3 mg/kg body weight. Plasma concentrations of morphine were followed for 4 h by a GC/MS technique. In most patients the peak plasma concentration was reached after 30 min; the mean peak plasma level of morphine was 18 ng/ml (range 8.5–57 ng/ml). The bioavailability of rectal morphine was determined in 6 patients, who received an i.m. injection of morphine at a second operation. The mean bioavailability of rectal morphine was 31% (range 12%–61%). None of the patients showed any clinical sign of respiratory depression, and there was no increase in end-tidal carbon dioxide tension measured in 5 patients operated under spinal block.

Notes: Summary The kinetics of ergotamine has been investigated in migrainous patients using a new, specific, sensitive HPLC assay (detection limit 100 pg/ml plasma). 10 patients were given ergotamine tartrate 0.5 mg i.v. and 5 of them received the same dose i.m. 2–3 weeks later. Blood samples were collected for up to 54 h following administration and the plasma concentration were analysed. After intravenous administration the plasma ergotamine declined rapidly, with an initial distribution half-life of 3 min followed by a mean terminal half-life of 1.86 h (range 90–155 min). The mean total plasma clearance was 11.0 ml kg−1 min−1, and the volume of distribution (Vdβ ) was 1847.6 ml kg−1. Individual t1/2β showed a positive linear correlation with the individual Vdβ . The intramuscular absorption of ergotamine was rapid and maximum plasma levels were usually obtained 10 min following administration. The biological availability was incomplete and variable at 46.6% (range 28.3–60.8%).

Notes: Summary The pharmacokinetic parameters of deacetyl metipranolol were determined after i.v. infusion of increasing doses (6–25 mg) in 17 normal volunteers. In a second cross-over trial, deacetyl metipranolol 10 and 20 mg were infused in a further 10 subjects, and in a third trial another 20 volunteers received metipranolol 40 mg orally. Metipranolol is very rapidly and completely deacetylated in man, so all pharmacokinetic data refer to deacetyl metipranolol, which was assayed by gas chromatography-mass spectrometry. The pharmacokinetic analysis was performed using a recently developed model, using a volume of distribution which is variable with time. The following data were obtained after oral administration: (mean values); lag-time 7.3 min; tmax 50 min, invasion half-life 6.3 min; elimination half-life 3 h; urinary excretion of unchanged drug approximately 4% of the dose. The experiments with infusion of increasing doses, as well as the cross-over study with 10 and 20 mg i.v., showed dose-linearity of the kinetics. The respective mean half-lives of elimination were 2.6, 2.9 and 2.8 h. The mean total, renal and extra-renal clearances amounted to 1237 ml/min, 149 ml/min and 1068 ml/min, respectively. The distribution coefficient was 3.5 l/kg, and protein binding amounted to 70% within the range of therapeutic concentrations. Absolute bioavailability was found to be approximately 50% by several different evaluation procedures. Thus, the pharmacokinetic profile of metipranolol shares features of both the lipophilic and the hydrophilic groups of β-blocking agents.

Notes: Summary 16 premature infants suffering from neonatal apnoea received orally an aqueous solution of theophylline 5 mg/kg bodyweight under fasting conditions and immediately before a milk feed. Bioavailability up to 7 h after administration was determined from the serum concentration-time course. The rate of absorption was significantly decreased if the drug was given with food; mean maximum serum concentrations were reached after 4.7 h instead of 1.6 h under fasting conditions. The area under the curve did not differ between the two patient groups which indicates that only the rate but not the amount of absorption was affected by food intake. The influence of feeding on the rate of absorption of theophylline by premature infants, which is more pronounced than in adults, can be related to particular functional factors in the gastrointestinal tract during the neonatal period.

Notes: Summary Acetylsalicylic acid (ASA) is a strong, irreversible inhibitor of platelet aggregation, but loses this activity following first-pass deacetylation to salicylic acid (SA). In order to compare the bioavailability of unchanged ASA from rapid- and slow-release formulations, the single-dose concentration profiles of ASA and SA were studied in healthy volunteers following intake of two different rapid-release (conventional and effervescent tablets) and three different slow-release (microencapsulated ASA in tablets and in capsules, and enteric-coated tablets) formulations of ASA, and of one slow-release formulation of sodium salicylate. Since anti-platelet therapy with ASA is often combined with dipyridamol, the influence of this drug was also examined. The concentrations of ASA and SA were measured by high-pressure liquid chromatography. While the bioavailability of SA from the 5 ASA formulations was essentially equal and similar to that of the salicylate formulation, the bioavailability and peak concentrations of ASA appeared to be the much greater after rapid-release than after slow-release formulations. Indeed, ASA was only rarely detected in systemic blood following intake of slow-release ASA. Co-administered dipyridamol did not significantly influence the kinetics of ASA or SA. It appears that rapid-release formulations of ASA should be prefered in anti-platelet therapy, either alone or in combination with dipyridamol.

Notes: Summary For drugs with a high hepatic clearance, bioavailability is low due to the so-called “first pass effect”. Prediction of the bioavailability for these drugs has been only lossely tested. It is proposed that by plotting the reciprocal of bioavailability versus the oral clearance, a straight line with intercept of unity and slope of reciprocal of hepatic blood flow should ensue. For lignocaine and verapamil, this relationship was found to be strong and gave good predictability, whereas for propranolol this relationship was weak and gave poor predictability. The proposed method may be of value in determining whether the low bioavailability of a drug is due to hepatic first pass metabolism.

Notes: Summary Plasma and urinary levels of chlorpheniramine (CPM) and its 2 demethylated metabolites were measured by HPLC after i.v. and oral dosing. In 5 mg (maleate) i.v. bolus studies in 2 subjects, plasma CPM levels were fitted to triexponential equations with terminal half-lives (t 1/2) of 23 and 22 h and area of 3.6 and 3.21/kg, respectively. Intravenous data predicted hepatic blood extraction ratios for the 2 subjects to be 0.06 and 0.07, respectively. Absolute bioavailability from oral solution (10 mg) was 59 and 34%, and from tablets (8 mg) 44 and 25%, respectively, indicating extensive gut first-pass metabolism. Mean t 1/2 from 7 oral fasting studies in 5 subjects was 28 h (19–43 h). Mean absorption lag time was 0.7 h (0.4–1.3 h), and mean peak time was 2.8 h (2–4 h). In 2 subjects, 6 mg solutions were given every 12 h for 9 doses; good correlation between single and multiple dose kinetics was found. Significant accumulation was demonstrated in simulation studies with frequent daily dosing. Estimated accumulation ratios vary from 4.1 to 9.4 (mean 6.5). The t 1/2 from urinary data (collected for 12 days) was consistent with plasma data. The above results suggest the need to reexamine the current practice of frequent daily dosing and the use of sustained or controlled release dosage forms of this drug. The possible cause of reduced plasma clearance of CPM in renal patients is discussed.

Notes: Summary The effect of food on the rate and extent of absorption of tolbutamide in diabetic patients was studied by varying the time of drug administration in relation to the ingestion of a standard meal. Serum levels of tolbutamide, insulin and glucose and related bioavailability parameters were compared following the administration of a single dose of tolbutamide 0.5 g to diabetic patients 30 min prior to and immediately before a standardized meal. A placebo dosage form was also administered to determine baseline glucose and insulin response to the meal. The 700 calorie standard meal was composed of 41% carbohydrate, 18% protein, and 41% fat. Administration of the drug with the meal resulted in a 6% (statistically significant) decrease in the extent of absorption, as determined by measurement of the area under the tolbutamide serum level-time curve from zero to infinity. Serum levels of tolbutamide were also significantly higher 0.5 h after drug administration when the drug was taken with the meal. Except for these two minor effects, no other differences between the drug treatments were observed in any other parameters of tolbutamide absorption or in the postprandial glucose and insulin serum levels. Therefore, the small differences found were judged to be clinically meaningless. These findings demonstrate that administration of tolbutamide 0.5 g tablets 30 min prior to or with a standard meal results in equivalent therapeutic actions.

Notes: Summary The kinetics of phenobarbital (PB) were evaluated in six normal subjects and six epileptic patients treated with phenytoin or carbamazepine. Each normal subject received three single doses of PB: PB-sodium 130 mg i.v. (IV), PB sodium 130 mg i.m. (IM), and PB acid 100 mg orally (PO), in random order at least one month apart. After IV PB distributive half-lives varied from 0.13 to 0.70 h, disposition half-lives were 75 to 126 h, steady state volume of distribution (Vss) was 0.54±0.03 l/kg, and clearance (CL) was 3.8±0.77 ml/h/kg. Absolute bioavailability of IM PB was 101±13%, of PO PB (corrected for dose) 100±11%. Peak serum PB concentrations were achieved from 2 to 8 h after IM administration, and from 0.5 to 4 h after PO administration. Epileptic patients exhibited similar PB kinetics: disposition half-lives were 77 to 128 h, Vss 0.61±0.05 l/kg, and Cl 3.9±0.76 ml/h/kg. Phenobarbital appears to represent an exception among antiepileptic drugs, in that pharmacokinetic data obtained in normals can reasonably be extrapolated to the epileptic population.

Notes: Summary The bioavailability of chloroquine in 7 healthy adult male volunteers was assessed with and without a standard breakfast. Chloroquine (600 mg base) was administered orally and timed blood samples were obtained for determination of plasma concentrations. The results suggest that the AUC and peak plasma levels were significantly higher when chloroquine was administered together with food, although the rate of absorption was not different. It appears, therefore, that food facilitates chloroquine absorption and the extent of absorption is significantly modified. Hence administration of chloroquine together with food appears to be an advisable clinical practice.

Notes: Summary 7-3H-phenylephrine was given to 15 volunteers by a short-infusionn=4) or p.o. (10 volunteers, 1 patient with porto-caval anastomosis). Analysis of serum for free3H-phenylephrine and fractionation of urinary radioactivity was performed by ion-exchange and thin-layer chromatography. As almost the same3H-activity was excreted in urine after i.v. and p.o. administration, 86% and 80% of the dose respectively, complete enteral absorption can be assumed. A considerable difference was seen in the fraction of free phenylephrine, i.v. 16% of the dose versus p.o. 2.6%, which suggested reduced bioavailability. This was confirmed by comparison of the areas under the serum curve, which showed a bioavailability factor of 0.38. The result for the patient with porto-caval anastomosis was comparable to that in the normal volunteers. The biological half-life of 2 to 3h was comparable to that of structurally related amines, as were the total clearance of 2 1/h, and the volume of distribution of 340l. Metabolism to phenolic conjugates mainly after oral ingestion, and tom-hydroxymandelic acid after i.v. injection, again demonstrated thatm-hydroxylated amines are predominantly conjugated during the “first-pass” metabolism.

Notes: Summary The elimination and bioavailability of two beta-blocking agents, propranolol and sotalol, were studied in 10 thyrotoxic patients, both before and after treatment with iodine-131. Each subject received in random order propranolol 160 mg and sotalol 160mg as single oral doses both while hyperthyroid and after euthyroidism had been achieved. The pharmacokinetics of sotalol was not affected by hyperthyroidism, whereas serum propranolol concentrations were significantly lower during hyperthyroidism than in the euthyroid state. During hyperthyroidism, the bioavailability of propranolol was significantly reduced (p〈0.05) and its clearance was increased (p〈0.005), whereas there was no difference in its serum t1/2. This indicates that the bioavailability of propranolol in hyperthyroidism is reduced by a mechanism which may depend on increased first-pass metabolism in the liver, or on an increased distribution volume of the drug. Both propranolol and sotalol caused a slight decrease in serum tri-iodothyronine concentration. As the effects of beta-blocking agents on the symptoms of hyperthyroidism are correlated with the serum concentration of the drugs, sotalol, with its long half-life and unaltered elimination in hyperthyroidism, has certain advantages over propranolol in the treatment of thyrotoxicosis.

Notes: Summary Two multiple-units controlled-release indomethacin capsule formulations containing enteric-coated pellets were bioequivalent to a standard capsule formulation (taken as the reference) in respect of extent of bioavailability in a crossover study with normal human subjects. However, drug absorption from the enteric-coated pellet formulations was slower, when compared to that from the standard reference capsule. The standard reference capsule released 85% of its drug content in vitro during 10 min at pH 6.5 and 98% during 1 h at pH 7.5. One enteric-coated pellet capsule formulation (I) released 77% during 1 h at pH 6.5 and the other (II) released 10% during 1 h at pH 6.5. Release of drug from each capsule of enteric-coated pellets was complete during 1 h at pH 7.5. Although differences in areas under the plasma indomethacin concentration-time curves were not significantly different, the peak plasma levels and the times of their occurrence indicated that the absorption rates of indomethacin decreased in the order, reference formulation 〉 pellet formulation I 〉 pellet formulation II, which was the same rank order as that of their dissolution rates in vitro. The data indicated that multiple units controlled-release formulations represent a reliable and reproducible source of indomethacin, which by avoiding extremes of local or systemic drug concentrations also should be better tolerated by individuals susceptible to unwanted gastrointestinal and centrally-mediated side-effects.

Notes: Summary Three different pivmecillinam preparations, a conventional 200 mg tablet (P tablet) and two new formulations containing respectively pivmecillinam 200 mg and 400 mg plus Avicel® (microcrystalline cellulose) as a disintegrator (PA tablet), were compared in vitro and in a gastroscopic study in 8 healthy volunteers. Disintegration of the PA tablet was significantly more rapid both in vitro and in the stomach. Following disintegration, the content of the PA tablet was spread over a larger area of the gastric mucosa (1088 mm2) than was observed with the P tablets (408 mm2). Three of the 8 volunteers taking the P tablet developed hyperaemia, interstitial bleeding or erosions of the mucosa of the stomach. No such reactions were seen with the PA tablets. Serum concentrations of mecillinam following ingestion of pivmecillinam tablets were determined in three groups of subjects; fasting volunteers, both supine and ambulant, and in ambulant subjects who took the preparation with a light meal. There was a tendency for the new PA tablets to produce a higher peak serum level as well as greater bioavailability of mecillinam. Administration of the PA tablets with a meal significantly increased the peak serum level and total bioavailability of the drug. On the basis of our observations we recommend adoption of the new PA tablet, because of its quick passage through the oesophagus and its more rapid and complete disintegration in the stomach.

Notes: Summary An high-pressure liquid chromatographic method was used to measure allopurinol and oxipurinol in plasma and urine in 6 healthy volunteers after a single intravenous or oral dose of allopurinol. The influence of coadministrated benzbromarone and hydrochlorothiazide on the pharmacokinetics of allopurinol and oxipurinol was also investigated. After intravenous injection of allopurinol 300 mg the plasma disappearance was biexponential, with a mean distribution half-life of 2.32±1.08 min $$(\bar x \pm SD)$$ and an elimination half-life of 47.8±10.6 min. The total clearance of allopurinol was 11.37±2.70 ml/min/kg, whereas its renal clearance was only 1.73±0.79 ml/min/kg. Oxipurinol disappeared monoexponentially from plasma ith a mean half-life of 12.2±2.6 h. Its renal clearance was 0.42±0.091 ml/min/kg. After oral administration of allopurinol 300 mg the peak plasma concentration of 2.1±0.6 µg/ml (1.5×10−5 M) was reached within 30 to 120 min. The peak level of oxipurinol of 5.8±1.5 µg/ml (3.8×10−5 M) was found within 2 to 5 h after intravenous and oral allopurinol. The bioavailability of oral allopurinol computed from plasma data was 90.4±8.7%. The total recovery from urine was 77% (allopurinol 8%, oxipurinol 69%) after oral and 88% after i.v. administration. It was concluded that about 10% of the oral dose was not absorbed and that 12% was eliminated by an unknown mechanism, presumably as riboside. The pharmacokinetics of allopurinol and oxipurinol were not significantly influenced by coadministration of benzbromarone or hydrochlorothiazide.

Notes: Summary Noscapine was administred to five healthy volunteers in a randomized crossover design, as an intravenous infusion of 66 mg, and as an oral 150 mg dose of either rapidly dissolving tablets or a tablet containing ion exchange resin-bound noscapine. After i.v. administration, the disposition of noscapine was bi-exponential with an elimination half-life of 2.6 h; the total plasma clearance was 22 ml/min/kg and the volume of distribution (Vdarea) was 4.7 l/kg. The absolute oral bioavailability was 30%, with a 3.6-fold interindividual variation. There was no pharmacokinetic evidence to support a prolonged action of the ion exchange resin tablet.

Notes: Summary The pharmacokinetics of bacmecillinam and pivmecillinam were studied in healthy fasting volunteers given tablets in a cross-over, randomized order. The mean (±SD) peak levels of plasma mecillinam were 1.43±0.34, 2.73±0.43, and 4.62±1.41 mg/l after bacmecillinam 100, 200, and 400 mg and 2.38±0.65 mg/l after pivmecillinam 400 mg. The corresponding areas under plasma Vs time curves (AUC) were 2.21±0.19, 3.99±0.63, and 7.74±1.38 mg·h·l−1 for bacmecillinam and 5.35±0.93 mg·h·l−1 for pivmecillinam. The elimination half-lives were 0.8–1.1h for bacmecillinam and 0.7h for pivmecillinam. The 12 h urinary recovery of unchanged mecillinam after the 400 mg doses was 41% for bacmecillinam and 30% for pivmecillinam. The 400 mg dose of bacmecillinam gave a significantly higher plasma peak (p〈0.001), AUC (p〈0.001) and urinary recovery (p〈0.001) than did pivmecillinam 400 mg. The plasma peaks appeared earlier and the rate of absorption was higher after bacmecillinam than after pivmecillinam (p〈0.05). In conclusion, bacmecillinam had a better bioavailability than pivmecillinam in the tablet formulations studied. The AUC increased linearly with increasing doses of bacmecillinam.

Notes: Summary The pharmacokinetics of terodiline HCl was studied in nine healthy volunteers given 12.5 mg i.v. and p.o. or 20 mg i.v. and 25 mg p.o. on two different occasions. The serum concentrations were measured by gas chromatography — mass spectrometry, using deuterated terodiline HCl as the internal standard. After i.v. administration the kinetics could be described by a two-compartment model with a mean distribution half life of 0.3 h and a mean elimination half life of 63 h. The serum clearance and apparent volume of distribution varied about 4-fold with mean values of 4.8 l/h and 417 l, respectively. After oral administration, the mean half life of absorption was 0.7 h and that of elimination 65 h. The absolute bioavailability varied between 64% and 105% with a mean of 92%. The long serum half life of terodiline should permit its once daily administration.

Notes: Summary Differences in the plasma concentrations of drugs after oral administration are caused by two main factors: variation in absorption ratios and in the distribution processes in the body. A new method for the dissection of both types of factors is discussed. The method uses a reference regression of the AUC-values to the corresponding values after intravenous infusion of graded doses. The reference regression is estimated from an appropriate trial. Deviation of the determined AUC-values from the regression curve afford an estimate of the residual variance due to varying distribution volumes or similar random biological effects. For the estimation of absorption ratios after oral administration the drug is given orally to another sample of subjects and their AUC-values are calculated. The deviation of these AUC values due to the above mentioned random effects are simulated using the residual variance of the reference regression, and are subtracted from the observed AUC-values. Then, the differences in the corresponding absorbed doses are transformed by inverting the reference regression. From these doses the empirical distribution function and statistical parameters (e.g. quantiles) are determined. The method has the advantage that no restrictive assumptions are required, such as first order processes, dose linearity, homogeneity of variance or normal distribution of absorption ratios. Its applicability to substances with qualitative differences in their pharmacokinetics is demonstrated by appropriate examples.

Notes: Summary Etilefrine undergoes considerable first-pass metabolism through conjugation in the gut wall. In six volunteers bioavailability was reduced to 35% for a fast release tablet and to 17% for a sustained release preparation. The addition fo dihydroergotamine (DHE) to the sustained release preparation surprisingly increased bioavailability to 61%. The plasma levels of the main metabolite formed during the passage through the gut wall indicated an increase in the rate of enteric absorption and therefore also in bioavailability by DHE. This might be due to the influence of DHE upon the vascular resistance of the vessels in the gut wall or a change in gastro-intestinal motility with a prolongation of drug contact time within the absorbing gut segment.

Notes: Summary In the case of liquid chromatography columns packed with small (3–5 μm) particles and utilized at relatively high flow rates, pressure pulses associated with the sample injection process (caused by a short-duration interruption in mobile phase flow to the column during injection) may result in a reduction of column lifetime. This detrimental effect can be eliminated by the utilization of specially constructed sample injection valves incorporating properly desinged bypass loops. The paper investigates in detail the effect of pressure pulses, describes the criteria and performance of the bypass systems, and documents column life time and performance.

Notes: Abstract Plasma concentrations of total and unbound sulfisoxazole were followed after single intravenous and oral doses of 1 g sulfisoxazole and during a 500-mg, four-time-a-day dosing regimen in six healthy males, using a specific high pressure liquid Chromatographic assay method. Saturable plasma protein binding was observed at total concentrations above 80–100 mg/liter. The clearance of sulfisoxazole was 18.7±3.9ml/min for total drug and 232±64ml/min for unbound drug. Renal elimination, on the average, accounted for 49% of the clearance of sulfisoxazole. The apparent volume of distribution for total drug was 10.9±2.0 liters and 136±36 liters for unbound drug, indicating that sulfisoxazole is primarily distributed extracellularly. Accumulation of N4-acetyl-sulfisoxazole during multiple dosing did not affect the disposition of sulfisoxazole. Adjusting for variable renal clearances between oral and intravenous administration and using the unbound plasma concentrations, the bioavailability for an oral dose of sulfisoxazole was found to be 0.95±0.04.

Notes: Abstract The bioavailability of phenytoin from rapid release capsule and oral solution formulations relative to that of a slow release capsule formulation was assessed in five patients who had participated in a three-way crossover study performed at steady state. The subjects then underwent dosage adjustment utilizing the slow release formulation, and estimates of their Michaelis-Menten parameters thus obtained were utilized in calculating the relative bioavailabilities. In addition, expected changes in steady-state plasma phenytoin concentrations were calculated assuming initial levels of 15 mg/liter, with increases and decreases in bioavailability of 10%. The consequences of such alterations in the extent of phenytoin absorption or average content of the dosage form may be clinically significant, particularly where the initial phenytoin level is equal to or greater than the patient's operative Km.

Notes: Abstract Current information on the pharmacokinetics of disopyramide is reviewed with emphasis on the implications for antiarrhythmic therapy. The absolute bioavailability, the disposition half-life, the plasma clearance, and the renal clearance for normal subjects and patients are discussed. Drug-drug interactions are discussed, and a new flexible intravenous dosing schedule is proposed.

Notes: Abstract The literature growth in pharmacokinetics and bioavailability between the years 1964 and 1980 is analyzed. During much of this period, the literature doubled approximately every 1.6 years. However, during the period 1978–1980, little or no growth was observed. During the period 1950–1967, the total chemical literature increased exponentially with a half-life of 8.28 years; between 1968 and 1980, the half-life was 12.4 years. Thus, the pharmacokinetic literature increased at a much more rapid pace than did the total chemical literature in general. The subject of growth is considered in a general context, particularly as influenced by psychological, sociological, political, and economic factors. It is concluded that while mathematical functions may adequately describe past literature trends, they have little if any utility in predicting future trends in specific research areas such as pharmacokinetics.

Notes: Abstract The pharmacokinetics of chloramphenicol (CAP) and total chloramphenicol succinate (CAPS) were studied in eight hospitalized adult patients with normal renal and hepatic function receiving intravenous chloramphenicol sodium succinate therapy. The steady-state peak concentrations of CAP (8.4–26.0 μg/ml) occurred at an average of 18.0 min (range 5.4–40.2) after cessation of the chloramphenicol sodium succinate infusion. Unhydrolyzed CAPS prodrug, representing 26.0±7.0% of the dose, was recovered unchanged in the urine indicating that the bioavailability of CAP from a dose of intravenous chloramphenicol succinate is not complete. A pharmacokinetic model was developed for simultaneous fitting of CAP and CAPS plasma concentration data. Pharmacokinetic parameters determined by simultaneous fitting were: V, 0.81±0.18 liters/kg; t1/2, 3.20 ±1.02 hr; CLB, 3.21±1.27 ml/min/kg for chloramphenicol; and V, 0.38±0.13 liters/kg; t1/2, 0.57±0.12hr; CLB, 7.72±1.87 ml/min/kg for total chloramphenicol succinate.

Notes: Abstract The pharmacokinetics and bioavailability of nitrazepam following intravenous, oral (tablet), and rectal (solution) administration were studied in seven healthy, young male volunteers. Nitrazepam plasma concentrations were determined by electron-capture GLC; pharmacokinetic evaluations were made by compartmental analysis (NONLIN) and compared with the results obtained by a less stringent modelling of the data. The plasma concentration-time profile was similar for all three routes of administration. Mean kinetic parameters as obtained by compartmental analysis of i.v. nitrazepam were: distribution half-life 17 min; volume of distribution after equilibrium 2.14 liters/kg; total plasma clearance 61.6 ml/min; elimination half-life 29.0 h. The mean protein unbound fraction of nitrazepam in plasma was 12.3% and the clearance of the unbound fraction was 506 ml/min. Absorption of oral nitrazepam started after the elapse of a lag time (mean value 12 min) and occurred as an apparent first-order process in all but one subject, with a mean absorption half-life of 16 min. Distribution and elimination half-lives were comparable with those following i.v. administration. Following rectal administration of the nitrazepam solution, rapid first-order absorption occurred with a mean lag time of 4 min and a mean absorption half-life of 9 min. Peak times (median 18 min) were significantly shorter than following oral administration (median 38 min), but there was little difference in peak concentrations. The distribution half-life was similar to i.v. and oral administration, but the elimination half-lives were longer with a mean value of 33.1 h. Following i.v. administration a good agreement was found between the results obtained by compartmental analysis using NONLIN and those obtained by a less stringent modelling of the data. Following oral and rectal administration, a good agreement between the two procedures was found for the elimination half-life; estimation of bioavailability, however, was higher by compartmental analysis. The mean bioavailability data showed that absorption is complete when nitrazepam is given orally and almost 20% lower when it is given rectally, but considerable interindividual differences were observed.

Notes: Abstract Seasonal and regional variations in the speciation, sediment-water partitioning, and dynamics of mercury (Hg) were studied at selected sites along the Hg-polluted Wabigoon River, and at unpolluted headwater and tributary sites, during April–September, 1979. ‘Dissolved’ and ‘particulate’ forms of Hg in the water were separated by continuous-flow centrifugation in the field. The Hg and other pollutants such as wood chips and salt had been discharged from a chlor-alkali plant and paper mill at Dryden, Ontario. Concentrations and loadings of particulate methyl mercury (CH3Hg+) and total particulate Hg (and loadings of total ‘dissolved’ Hg) were greatest during the spring flood (April-May) owing to accelerated resuspension and transport of sediments. Concentrations of ‘dissolved’ CH3Hg+, however, were highest in the summer (July–September), probably reflecting stimulation of microbial methylating activity by elevated temperatures, together with factors such as reduced levels of metal-scavenging particulates and minimal dilution by runoff. Total dissolved Hg concentrations were relatively high in September at polluted sites only, possibly because of desorption from sediments due to elevated concentrations of Cl− ions. Loadings of dissolved CH3Hg+ tended to be high in the summer but were generally depressed (suggesting sorption by suspended particles) during the major spring-flood episode in May. During July–August dissolved CH3Hg+ was a function of total dissolved Hg, suggesting rapid biomethylation of desorbed inorganic Hg; but in general dissolved and suspended CH3Hg+ levels depended on environmental variables and were unrelated to total Hg concentrations. In the summer only, total dissolved Hg was a function of dissolved Cl−. Hg species in particulates were associated with sulfides, hydrated Fe and Mn oxides, organic matter (notably high molecular weight humic and humic-Fe components), and selenium (Se); but CH3Hg+ and total Hg differed in their specific preferences for binding agents, implying that binding sites discriminate between CH3Hg+ and Hg2+ ions. CH3Hg+ was associated with sulfide and (in the spring only) with Fe oxides, whereas total Hg was associated with organic matter and Se and with DTPA- and NaOH-extractable Fe in the spring but with Mn oxide and NaOH-extractable organics in the summer. Sulfides were most abundant in May, indicating that they were eroded from bottom sediments, but Fe and Mn oxides were most abundant in the summer, probably owing to activities of filamentous iron bacteria and other micro-organisms. Particulate Hg was 98–100% nonextractable by mild solvents such as Ca acetate, CaCl2, dilute acetic acid, and (at polluted sites only) DTPA solutions, suggesting that the particulate Hg mobilized in the spring may not be readily available to organisms; association with Se and high molecular weight humic matter also supports this hypothesis. Hg probably becomes more bio-available in the summer, as suggested by the upsurge in dissolved CH3Hg+ and total dissolved Hg levels, and by increases in the solubility of particulate Hg in acetic acid, DTPA, H2O2, and NaOH solutions, as well as an increase in the relative importance of lower molecular weight fractions of NaOH-extractable Hg (in September). Regional variations in Hg speciation and partitioning reflected a gradient in sediment composition from wood chips near Dryden to silt-clay mud further downstream. Hg in silt-clay mud relatively far (〉 35 km) downstream from the source of pollution or in unpolluted areas appeared to be more readily solubilized by Cl− ions or chelators such as DTPA, more readily methylated (as indicated by downstream increases in dissolved CH3Hg+ levels and CH3Hg+/total Hg ratios), and was to a greater degree organically bound (H2O2-extractable), and thus was probably more bio-available, than Hg in wood-chip deposits. Possible explanations include weaker binding of Hg by the mud, the more finely divided state of the mud, and improved microbial growth at lower concentrations of toxic pollutants. Owing to enrichment in sulfides and Fe oxides, resuspended wood-chip sediments were especially efficient scavengers of CH3Hg+. The results indicate that in any pollution abatement plan aimed at lowering the Hg levels in the biota of lakes fed by the Wabigoon River, immobilization, removal, or detoxification of dissolved as well as particulate forms of Hg in the river would probably have to be considered. Possibly, Hg species could be ‘scrubbed’ from the river water by increasing the suspended load and by sedimentation and treatment with Hg-binding agents in special receiving basins.

Notes: Abstract Seasonal and regional variations in the speciation, sediment-water partitioning, and dynamics of mercury (Hg) were studied at selected sites along the Hg-polluted Wabigoon River, and at unpolluted headwater and tributary sites, during April–September, 1979. ‘Dissolved’ and ‘particulate’ forms of Hg in the water were separated by continuous-flow centrifugation in the field. The Hg and other pollutants such as wood chips and salt had been discharged from a chlor-alkali plant and paper mill at Dryden, Ontario. Concentrations and loadings of particulate methyl mercury (CH3Hg+) and total particulate Hg (and loadings of total ‘dissolved’ Hg) were greatest during the spring flood (April-May) owing to accelerated resuspension and transport of sediments. Concentrations of ‘dissolved’ CH3Hg+, however, were highest in the summer (July–September), probably reflecting stimulation of microbial methylating activity by elevated temperatures, together with factors such as reduced levels of metal-scavenging particulates and minimal dilution by runoff. Total dissolved Hg concentrations were relatively high in September at polluted sites only, possibly because of desorption from sediments due to elevated concentrations of Cl− ions. Loadings of dissolved CH3Hg+ tended to be high in the summer but were generally depressed (suggesting sorption by suspended particles) during the major spring-flood episode in May. During July–August dissolved CH3Hg+ was a function of total dissolved Hg, suggesting rapid biomethylation of desorbed inorganic Hg; but in general dissolved and suspended CH3Hg+ levels depended on environmental variables and were unrelated to total Hg concentrations. In the summer only, total dissolved Hg was a function of dissolved Cl−. Hg species in particulates were associated with sulfides, hydrated Fe and Mn oxides, organic matter (notably high molecular weight humic and humic-Fe components), and selenium (Se); but CH3Hg+ and total Hg differed in their specific preferences for binding agents, implying that binding sites discriminate between CH3Hg+ and Hg2+ ions. CH3Hg+ was associated with sulfide and (in the spring only) with Fe oxides, whereas total Hg was associated with organic matter and Se and with DTPA- and NaOH-extractable Fe in the spring but with Mn oxide and NaOH-extractable organics in the summer. Sulfides were most abundant in May, indicating that they were eroded from bottom sediments, but Fe and Mn oxides were most abundant in the summer, probably owing to activities of filamentous iron bacteria and other micro-organisms. Particulate Hg was 98–100% nonextractable by mild solvents such as Ca acetate, CaCl2, dilute acetic acid, and (at polluted sites only) DTPA solutions, suggesting that the particulate Hg mobilized in the spring may not be readily available to organisms; association with Se and high molecular weight humic matter also supports this hypothesis. Hg probably becomes more bio-available in the summer, as suggested by the upsurge in dissolved CH3Hg+ and total dissolved Hg levels, and by increases in the solubility of particulate Hg in acetic acid, DTPA, H2O2, and NaOH solutions, as well as an increase in the relative importance of lower molecular weight fractions of NaOH-extractable Hg (in September). Regional variations in Hg speciation and partitioning reflected a gradient in sediment composition from wood chips near Dryden to silt-clay mud further downstream. Hg in silt-clay mud relatively far (〉 35 km) downstream from the source of pollution or in unpolluted areas appeared to be more readily solubilized by Cl− ions or chelators such as DTPA, more readily methylated (as indicated by downstream increases in dissolved CH3Hg+ levels and CH3Hg+/total Hg ratios), and was to a greater degree organically bound (H2O2-extractable), and thus was probably more bio-available, than Hg in wood-chip deposits. Possible explanations include weaker binding of Hg by the mud, the more finely divided state of the mud, and improved microbial growth at lower concentrations of toxic pollutants. Owing to enrichment in sulfides and Fe oxides, resuspended wood-chip sediments were especially efficient scavengers of CH3Hg+. The results indicate that in any pollution abatement plan aimed at lowering the Hg levels in the biota of lakes fed by the Wabigoon River, immobilization, removal, or detoxification of dissolved as well as particulate forms of Hg in the river would probably have to be considered. Possibly, Hg species could be ‘scrubbed’ from the river water by increasing the suspended load and by sedimentation and treatment with Hg-binding agents in special receiving basins.

Notes: Abstract Control of phytoplankton production in the Great Lakes can be achieved most efficiently by limiting inputs of biologically available P. We report the results of studies performed to characterize the chemical forms and availability of particulate P in wastewater and tributaries which enter the lower Lakes, the eroding bluffs which border Lake Erie, and bottom samples from the near-shore of western Lake Erie. Rates of release of available P were estimated from a simple first-order model of the process, as observed during algal bioassays. Available P in wastewater samples, as a fraction of total particulate P, was affected minimally by wastewater treatment, including chemical precipitation and filtration; it correlated well with levels of total particulate P. Available particulate P levels in fluvial suspended sediments showed regional uniformity, but appeared to be strongly dependent on levels of both NaOH-P and CDB-P. Rates of release of available P decreased during wastewater treatment to values which were similar in magnitude to those observed for fluvial sediments. Release rates, however, were not related to any of the particulate P fractions which were measured. Analysis of the bluff and bottom samples indicated that P availability in the former was negligible, but the latter contained levels which approached those of wastewater particulates, though available P was released from the bottom sediments at relatively low rates.

Notes: Abstract Control of phytoplankton production in the Great Lakes can be achieved most efficiently by limiting inputs of biologically available P. We report the results of studies performed to characterize the chemical forms and availability of particulate P in wastewater and tributaries which enter the lower Lakes, the eroding bluffs which border Lake Erie, and bottom samples from the near-shore of western Lake Erie. Rates of release of available P were estimated from a simple first-order model of the process, as observed during algal bioassays. Available P in wastewater samples, as a fraction of total particulate P, was affected minimally by wastewater treatment, including chemical precipitation and filtration; it correlated well with levels of total particulate P. Available particulate P levels in fluvial suspended sediments showed regional uniformity, but appeared to be strongly dependent on levels of both NaOH-P and CDB-P. Rates of release of available P decreased during wastewater treatment to values which were similar in magnitude to those observed for fluvial sediments. Release rates, however, were not related to any of the particulate P fractions which were measured. Analysis of the bluff and bottom samples indicated that P availability in the former was negligible, but the latter contained levels which approached those of wastewater particulates, though available P was released from the bottom sediments at relatively low rates.

Notes: Abstract Seasonal and regional variations in the speciation, sediment-water partitioning, and dynamics of mercury (Hg) were studied at selected sites along the Hg-polluted Wabigoon River, and at unpolluted headwater and tributary sites, during April–September, 1979. ‘Dissolved’ and ‘particulate’ forms of Hg in the water were separated by continuous-flow centrifugation in the field. The Hg and other pollutants such as wood chips and salt had been discharged from a chlor-alkali plant and paper mill at Dryden, Ontario. Concentrations and loadings of particulate methyl mercury (CH3Hg+) and total particulate Hg (and loadings of total ‘dissolved’ Hg) were greatest during the spring flood (April-May) owing to accelerated resuspension and transport of sediments. Concentrations of ‘dissolved’ CH3Hg+, however, were highest in the summer (July–September), probably reflecting stimulation of microbial methylating activity by elevated temperatures, together with factors such as reduced levels of metal-scavenging particulates and minimal dilution by runoff. Total dissolved Hg concentrations were relatively high in September at polluted sites only, possibly because of desorption from sediments due to elevated concentrations of Cl− ions. Loadings of dissolved CH3Hg+ tended to be high in the summer but were generally depressed (suggesting sorption by suspended particles) during the major spring-flood episode in May. During July–August dissolved CH3Hg+ was a function of total dissolved Hg, suggesting rapid biomethylation of desorbed inorganic Hg; but in general dissolved and suspended CH3Hg+ levels depended on environmental variables and were unrelated to total Hg concentrations. In the summer only, total dissolved Hg was a function of dissolved Cl−. Hg species in particulates were associated with sulfides, hydrated Fe and Mn oxides, organic matter (notably high molecular weight humic and humic-Fe components), and selenium (Se); but CH3Hg+ and total Hg differed in their specific preferences for binding agents, implying that binding sites discriminate between CH3Hg+ and Hg2+ ions. CH3Hg+ was associated with sulfide and (in the spring only) with Fe oxides, whereas total Hg was associated with organic matter and Se and with DTPA- and NaOH-extractable Fe in the spring but with Mn oxide and NaOH-extractable organics in the summer. Sulfides were most abundant in May, indicating that they were eroded from bottom sediments, but Fe and Mn oxides were most abundant in the summer, probably owing to activities of filamentous iron bacteria and other micro-organisms. Particulate Hg was 98–100% nonextractable by mild solvents such as Ca acetate, CaCl2, dilute acetic acid, and (at polluted sites only) DTPA solutions, suggesting that the particulate Hg mobilized in the spring may not be readily available to organisms; association with Se and high molecular weight humic matter also supports this hypothesis. Hg probably becomes more bio-available in the summer, as suggested by the upsurge in dissolved CH3Hg+ and total dissolved Hg levels, and by increases in the solubility of particulate Hg in acetic acid, DTPA, H2O2, and NaOH solutions, as well as an increase in the relative importance of lower molecular weight fractions of NaOH-extractable Hg (in September). Regional variations in Hg speciation and partitioning reflected a gradient in sediment composition from wood chips near Dryden to silt-clay mud further downstream. Hg in silt-clay mud relatively far (〉 35 km) downstream from the source of pollution or in unpolluted areas appeared to be more readily solubilized by Cl− ions or chelators such as DTPA, more readily methylated (as indicated by downstream increases in dissolved CH3Hg+ levels and CH3Hg+/total Hg ratios), and was to a greater degree organically bound (H2O2-extractable), and thus was probably more bio-available, than Hg in wood-chip deposits. Possible explanations include weaker binding of Hg by the mud, the more finely divided state of the mud, and improved microbial growth at lower concentrations of toxic pollutants. Owing to enrichment in sulfides and Fe oxides, resuspended wood-chip sediments were especially efficient scavengers of CH3Hg+. The results indicate that in any pollution abatement plan aimed at lowering the Hg levels in the biota of lakes fed by the Wabigoon River, immobilization, removal, or detoxification of dissolved as well as particulate forms of Hg in the river would probably have to be considered. Possibly, Hg species could be ‘scrubbed’ from the river water by increasing the suspended load and by sedimentation and treatment with Hg-binding agents in special receiving basins.

Notes: Summary The pattern of follicular development during the estrous cycles of aged rats was examined and compared with that of mature rats. In both, preovulatory follicles are derived from a select group of small pre-Graafian follicles which begin to develop at estrus and reach the preovulatory size by the morning of proestrus, but the rate of growth, as judged by an increase in the percentage of granulosa cells incorporating 3H-thymidine, is accelerated in the follicles of aged rats. A second mechanism, which accounts for preovulatory follicles in aged rats, involves the rescue from atresia of pre-Graafian and preovulatory follicles. The existence of this mechanism is supported by the observation that at metestrus in aged rats virtually all follicles, regardless of their state of atresia, possess a high percentage of granulosa cells incorporating 3H-thymidine, indicating that the follicles are growing rapidly. However, some of these rapidly growing follicles show signs of atresia such as pyknotic nuclei within their granulosa cell layers. Since follicles in the initial stage of atresia contain defective oocytes (Peluso et al. 1979b), their rescue and development into preovulatory follicles would result in the ovulation of defective oocytes, a fact which accounts in part of the lower fertility in these older animals.

Notes: Summary The postnatal development of microglial cells was investigated in the neonatal rat brain by use of light- and electron microscopy, including enzyme-histochemical techniques. Microglial cells were selectively stained by demonstration of their nucleoside diphosphatase (NDPase) activity and classified into three types: 1) In the early postnatal period, “primitive microglial cells” showing scantily ramified processes were found in the cerebral cortex, the hippocampal formation, and the hypothalamus. During the course of the first postnatal week the processes of this cell type developed gradually and the cells were transformed into typical ramified microglial cells, called “resting microglial cells”. 2) “Amoeboid microglial cells” showing typical features of macrophages were characteristic of the cerebral white matter. 3) “Round microglial cells” possessing a round soma and few pseudopodia but no characteristic processes occurred in large numbers in the sub ventricular zone of the lateral ventricle and as single elements in the vicinity of blood vessels. Histochemically, thiamine pyrophosphatase (TPPase) was demonstrated only in the fully developed, ramified microglial cells (“resting microglial cells”), which could be readily observed in the central nervous tissue from the age of 14 day. “Round and amoeboid microglial cells” did not show TPPase activity and disappeared after 14 days of postnatal life. By use of electron microscopy, in neonatal rats NDPase activity was apparent in the plasma membrane of the three types of microglial cells (“primitive, round, and amoeboid” types). They showed basically similar submicroscopic characteristics, i.e., well-developed Golgi apparatus, long strands of roughsurfaced endoplasmic reticulum, single dense bodies and vacuoles, and numerous ribosomes. “Amoeboid microglial cells” were characterized by their well-developed cytoplasmic vacuoles and phagocytic inclusion bodies. The present study strongly suggests a mesodermal origin for these microglial elements.

Notes: Summary Five common tricyclic antidepressants are separated on a silica column using an aqueous eluent. The tricyclics are concentrated from serum using an adsorption procedure on a solid phase Sep PakTM C18 cartridge. The method appears to be adequate for monitoring therapeutic levels of these drugs.

Notes: Summary Four different methods of calculation of retention in ternary mobile phases were compared and it was found that the simple calculation based only on two values of the capacity factors (one for each binary system that composes the ternary mobile phase) provides the accuracy of prediction that is, at least, comparable to the other methods of calculation that require a large number of preliminary experiments. The deviations from ideal behaviour in ternary solvent mixtures are discussed; some sources of errors can be avoided, at least partially, using binary systems of adequate compositions for preparation of ternary mobile phases. Several examples show the comparison of calculated and experimental selectivities in ternary solvent systems. A simple calculation can be used for rapid selection of an adequate ternary (or more complex) mobile phase, if it is necessary to achieve the separation of a given sample mixture.

Notes: Summary The efficiency of high-performance liquid chromatography columns packed with 5 μm and 3 μm reverse-phase octadecylsilane packing materials has been evaluated using four test mixtures. Both stopped flow and valve injection have been used and typical efficiencies are given.

Notes: Summary It is shown that sensitized room-temperature phosphorescence in liquid solutions can be successfully applied as a detection method in continuous flow and chromatographic systems. The limits of detection for a series of halogenated naphthalenes and biphenyls obtained by plug injection are roughly the same as those in batch experiments — in the subnanogram region. Liquid chromatograms of mixtures of PCBs, PCNs and polychlorodibenzofurans are given.

Notes: Summary A specific HPLC method has been developed for the trace analysis of lower chlorinated aromatic compounds. The method consists of an on-line preconcentration and a post-column reaction step. On-line preconcentration of mono- and dichlorophenols from aqueous samples has been performed using PRP1, a divinylbenzene-styrene copolymeric sorbent as packing material for both pre-and analytical column. Enrichment factors of over 300 were obtained compared to regular (100 μl) loop injections, even for the highly polar monochlorophenols. After reversed-phase separation, post-column photochemical dechlorination is carried out directly in the eluent stream, using a photochemical reactor. Upon dechlorination, fluorescent products are formed, which can be detected selectively. The resulting fluorescence signal shows a linear response to the quantity of solute present over 2 to 3 orders of magnitude (correlation coefficients: 0.990–0.98). For the mono- and dichlorophenols, the detection limit of the photoconversion method is in the lower nanogram range. The method is especially suitable for the analysis of complex matrices such as effluent water samples or biological fluids containing traces of the polar chloroaromatic compounds together with large amounts of other constituents, which interfere when using more general methods of detection like UV absorption. The potential of this technique for automation has been demonstrated by using a microprocessor-controlled column switching unit.

Notes: Summary Compounds without own detector response can be detected and quantified by a UV-absorbing or fluorescent, ion-pairing probe in reversed-phase liquid-solid systems. Charged as well as uncharged samples give response by affecting the distribution of the probe between the phases. The probe is usually a hydrophobic organic ion. It is included in the mobile phase, which also contains a counter ion and an ion of the same charge as the probe, both fairly hydrophilic. The choice of properties and concentrations of the hydrophilic ions is essential for the response since too low and too high distribution of the mobile phase ions to the adsorbent will make the system insensitive. It is with UV-absorbing probes possible to get a response corresponding to a molar absorptivity of more than 3000 for ionic samples. Fluorescent probes have given 3–5 times higher sensitivity. Uncharged compounds give as a rule lower response. A combination of two UV-absorbing probes with different charge gives a considerable improvement of the response for samples with low retention. A simplified theoretical model for the detector response is suggested on the basis of detailed studies of the concentration changes in the eluted mobile phase.

Notes: Summary A new photochemical reaction method for the on-line fluorimetric detection of natural vitamin K homologs was developed. In addition to its high sensitivity, the method features an interesting selectivity for the determination of these compounds in a complex matrix such as human serum. The formation of different photolysis products as a function of reaction conditions, and the optimalisation of a segmented flow reaction system will be discussed. The results indicate that the method is 4 times more sensitive than UV detection, and at least as selective as fluorescence detection after post-column chemical reduction.

Notes: Summary The mass detector was investigated as an alternative to RI detection for carbohydrate and lipid analyses with high pressure liquid chromatography. The instrument was found to be more sensitive than most RI detectors and, as gradient elution could also be carried out, the resulting chromatograms showed improved resolution.

Notes: Summary The retention behaviour of nucleobases and nucleosides on unmodified silica with dichloromethane-methanol-water mixtures has been systematically investigated. The degree and order of retention can be varied over a wide range by changing the pH, the type and concentration of the acidic additives and by the methanol and water content of the mobile phase. The retention process cannot be considered as simple adsorption but rather as a very complex mixed distribution process of adsorption and absorption including the involvement of ion-pair formation. Further data on the effect of the type of silica (source of supply) on retention behaviour, column efficiency and column reproducibility are reported. The practical application of unmodified silica for the separation of nucleobases and nucleosides is demonstrated by the analysis of a hydrolysate of calf thymus DNA.

Notes: Summary A reaction detector with non-segmented flow in open tubes as reaction track is described. To minimize peak broadening the open tubes are arranged in a three dimensional coiled structure by knitting. At low volume flow rates the h-values in these open tubes are independent of flow rate and are significantly lower than predicted by theory. The applicability of this reactor for classical colorimetric detection with corrosive reagents is shown.

Notes: Summary This paper is concerned with our investigation into the effects of temperature on the liquid chromatography absorbance baseline. Measurements were made to quantify the effects in terms of wavelength and solvents. Five solvents commonly used in HPLC were investigated, methanol, propanol, hexane, heptane and acetonitrile. The alcoholic solvents were found to have a high absorbance sensitivity to temperature at wavelengths around their ‘UV cut off’ points. Possible mechanisms for this effect are discussed and include solvent impurity, density changes and inter-molecular bonding.

Notes: Summary The paper presents an overview of the evolution of open tubular (capillary) columns introduced 25 years ago by M.J.E. Golay. This evolution is divided into five stages: the formulation of the theory and the basic work laying down the foundations for future work; learning the influence of the variables on column performance; development of columns with increased surface area; development of glass and fused-silica columns; and, finally, the introduction of open tubular columns in liquid chromatography. In each stage, the pioneering activities are summarized and the most important references given.

Notes: Summary The reaction of a newly developed fluoregenic reagent, 7-fluoro-4-nitrobenzo-2-oxa-1,3-diazole(NBD-F), with amino acids and biogenic amines was investigated. NBD-F was reactive to both primary and secondary amines including amino acids and biogenic amines such as catecholamines. The amino acids were reacted with the reagent, separated by high-performance liquid chromatography on μ-Bondapak C18 and detected at 10 to 100 fmol level. A few μg of protein hydrolysates, rabbit pyruvate kinase M1, rabbit aldolase A and papain, were adequate for the amino acids quantitation. An automatic amino acid analyzer with fluorometric detection by the post-column derivatization with NBD-F enabled the amino acid profile analysis in blood samples present in a paper disc of 3 mm diameter.

Notes: Summary Extracts of insect, mite, fungal and mammalian origin, known to provoke an allergic response in man, have been examined and fractionated by high-performance size exclusion chromatography. The mild conditions employed for separation have permitted direct immunological assessment of the fractions obtained, and recognition of the biologically important components. These are being isolated for further study. This chromatographic procedure has proved to be of value for assessing different extracts from the same source material as potential reference preparations. It can also be employed as a sensitive method for identification of extracts derived from a variety of sources.

Notes: Summary N-Protected aminoacids have been chemically bonded to aminopropylated silica gel, and the resulting chiral phases have been applied to the LG resolution of racemic mixtures of polar compounds. Best results were obtained with surfacebound N-formyl-isoleucine and N-formylvaline, with which baseline resolutions of a range of aminoacid and aminoalcohol derivatives were achieved.

Notes: Summary An improved interface for the coupling of a liquid chromatograph (LC) and an electron-capture detector (ECD) allows the use of the LC-ECD combination for the determination of various types of chlorinated aromatic compounds. The addition of up to 6% of dioxan to the hexane used as mobile phase can be tolerated without serious deterioration of detector performance. Derivatization of polar compounds such as anilines, phenylurea herbicides, chlorophenols and hydroxylated polychlorobiphenyls with fluorine-and non-fluorine-containing acid anhydrides is a useful tool to extend the application range of LC-ECD. As an example, the analysis of a spiked soil sample is discussed.

Notes: Summary A flow-through two electrode wall-jet cell with a platinum measuring electrode and a cell volume of 20 nl has been designed and evaluated. It has been used to detect phenols by reversed phase liquid chromatography using short micro bore columns. The linear dynamic range between the measured current and the concentration is greater than 103 (1.5×10−7−5×10−4 mol/l) and the minimum analyzable amount was found to be 10 pg for pyrocatechol. A negligible broadening in the detector permits the use of micro columns down to 0.5 mm internal diameter, packed with 5 μm particles, without any substantial distortion of eluted zones.

Notes: Summary A silica of 14 nm pore size was reacted according to two procedures with the following silanes: γ-aminopropyltriethoxysilane (1), N-aminoethyl-N′-aminopropyltrimethoxysilane (2), N-aminoethyl-N′-aminoethyl-N″-aminopropyltrimethoxysilane (3), N-(3-triethoxysilanepropyl)-N, N-diacetic dimethylester (4), N-(3-trisodiumsilanolatepropyl)-N, N-diacetic acid disodium salt (5) and N-(3-trisodiumsilanolatepropyl)-ethylenediamine-N, N′, N′-triacetic acid trisodium salt (6). The reaction of silanes 1–4 with the silica was carried out under anhydrous conditions (procedure A). Silanes 1–6 were subjected to surface modification under essentially hydrous conditions applying a two-step procedure (procedure B). Procedure B is much simpler to perform and allows binding of high and definite amounts of silane onto the surface. Complexation of all bonded phases with Cu2+ was examined by measuring the sorption isotherms at constant pH. The pH-dependence of Cu2+ uptake at constant Cu2+ concentration was evaluated. Tests on the chemical and chromatographic stability of the packings showed that only the silicas with bonded iminodiacetate and ethylenediaminetriacetate groups remain hydrolytically stable. Retention of aliphatic and α-amino acids on these two packings was found to be mainly controlled by the pH and the ionic strength of the eluent.

Notes: Summary A novel form of liquid chromatography is described which involves the use of multiphase eluents, such as emulsions, for the separation of biopolymers and certain microorganisms.

Notes: Summary The effect of nonpolar (hexane, decane, benzene) and polar (water, isopropanol, water+isopropanol) eluents on separation of aromatic hydrocarbons and their derivatives in liquid-adsorption chromatography on hydroxylated silica gel columns has been studied. The work has revealed that the traditional application areas of hydroxylated silica gel in liquid chromatography may be extended. In particular it has been shown that, first, polar compounds may be separated on hydroxylated silica gel with nonpolar eluents at high temperatures (100–150°C) and that polar and nonpolar compounds may be separated using pure water as eluent. The effect of adsorbed moisture on retention behaviour of silica gel with nonpolar eluents has also been investigated.

Notes: Summary A detailed study of isocratic and gradient elution separations of lanthanides has been carried out. Analyses of industrially and scientifically interesting products such as luminescent phosphors have been carried out by gradient elution with DL-2-hydroxyisobutyric acid. The determination of small amounts of terbium in gadolinium oxide sulphide phosphors is described in which an HCl solution was eluted through a stainless steel column packed with microparticulate silica, with bonded cation-exchange groups. Complete separation of gadolinium and terbium is achieved. Detection is with a variable wavelength detector following post-column complex formation with 4-(2-pyridylazo)-resorcinol monosodium salt. Results obtained on test solutions show good reproducibility and sensitivity and the method may be considered sufficiently reliable to be used in routine quality control procedures.

Notes: Summary For the separation of carbohydrates by liquid chromatography, utilizing the intermolecular interaction with amino groups on the surface of the adsorbent, two types of adsorbents were used: silica gel modified by adding to the eluent a diamine, and a chemically bonded phase prepared by the reaction of γ-aminopropyl triethoxy silane with silica gel. Mono- di- and trisaccharides could be separated on silica gel modified by the adsorption of piperazin and ethylene diamine from the eluent. The separation capacity of columns packed with silica gel with bonded NH2 groups and with silica gel having diamines adsorbed on its surface is similar. The retention volumes of 15 carbohydrates were measured on columns containing silica gel with hydroxylated surface and on silica gel modified with piperazin from the eluent consisting of acetone-water. Comparison of acetone-water and acetonitrile-water as the eluent showed that the former is also suitable for the analysis of carbohydrates.

Notes: Summary Reaction detectors are considerably enhancing the application potential of HPLC particularly when dealing with trace analysis in complex matrices. In principle one distinguishes between segmented and non-segmented reactor system. The theory of band broadening of these designs and some criteria for the choice of the proper type are brieffly discussed. It seems that with a proper construction of the phase separators segmented systems have a wider range of applicability. Applications of reaction detectors in the areas of pharmaceutical and environmental analysis are discussed.

Notes: Summary Resolution (Rs) is a function of three factors: efficiency, retentivity and selectivity. A procedure for optimization of mobile phase selectivity was demonstrated using a reversed-phase (RP) separation of compounds with various functionalities. In this paper, the same procedure is used with normal bonded-phase (NP) chromatography to develop a separation of structurally similar steroids. The optimization of mobile-phase selectivity provided increased problem-solving capability and decreased the analysis time.

Notes: Summary The responding signals of eluted components can be enhanced by using high performance small volume columns with long lengths generating more plates than required for a separation with a preset resolution in combination with instruments which show very small external bandbroadening. The excess number of plates is then consequently erased by maximizing the injection volume of the sample until the preset resolution is reached. Equations, describing the dilution, relative signal enhancement and maximum injection volume as a function of the process parameters are derived and experimentally verified. Theory and experiment are in agreement only if variances of response functions are calculated as their second normalized central moments.

Notes: Summary The noradrenergic terminals in the substantia gelatinosa of the dorsal horn of the cervical spinal cord of the rat were investigated by means of the histofluorescence technique and electron-microscopic cytochemistry using the glyoxylic acid-KMnO4 fixation technique. In accordance with the topographical distribution of fluorescent catecholaminergic fibers, noradrenergic terminals containing small granular vesicles were frequently observed electron microscopically in the outer layer of the substantia gelatinosa. These terminals were most frequently found to appose without showing typical synaptic features, small-caliber dendrites, spine apparatus, and rarely, large caliber dendrites. Only in a few cases, the noradrenergic terminals exhibited typical synaptic contacts with dendritic elements of small size. In addition, noradrenergic terminals apposed non-noradrenergic terminals containing small agranular vesicles. In rats bearing surgical lesions of the dorsal roots, no noradrenergic terminal were found in contact with the degenerated axon terminals in the substantia gelatinosa. These findings suggest that the noradrenergic afferents to the substantia gelatinosa may exert their influence on sensory transmission via dorsal horn cells.

Notes: Summary In a study of the effect of pinealectomy on thyroid C-cell number, 8 animals out of 66 were found to have thymic tissue in close association with the thyroid. Cells containing immunoreactive calcitonin were found in all of the thyroids but in only one of the 8 pieces of thymus. These cells found in a piece of thymic tissue associated with the right thyroid lobe were located immediately under the capsule and did not form or associate with follicles. Unlike the other animals the rat with thymic calcitonin cells had an unequal distribution of C-cells between the left and right thyroid lobes, but the total number of thyroidal C-cells was the same as that of the other rats. Since the thymus proper was not examined in these 66 animals, ten additional rats were taken for such a study. Thyroid-associated thymic tissue was found in three of these, but none of these thymi showed any immunoreactive cells.

Notes: Summary The autonomic innervation of rat jugular vein was studied using glyoxylic acid fluorescence and acetylcholinesterase histochemical methods. The rat jugular vein is provided with both adrenergic and cholinergic nerve fibers organized in plexuses located at the adventitial-medial border. The existence of these nerve plexuses does not seem to support biochemical findings that suggest a lack of innervation in the rat jugular vein and which propose this blood vessel as a model for the analysis of drug-smooth muscle cell interaction without the interference of neuronal uptake mechanisms.

Notes: Summary Electron-microscopic survey of selectively stained microglial cells in the cerebral cortex of the rat reveals that the processes of this cell type often encircle axo-dendritic synapses. Enzyme-histochemical methods for thiamine pyrophosphatase (TPPase) or nucleoside diphosphatase (NDPase) were used for the selective marking of the microglial cells; TPPase and NDPase activities were observed in the plasma membrane of microglial cells. The synapses encircled by microglial processes displayed presynaptic structures containing round clear vesicles (50 nm in diameter) and a prominent thickening of the postsynaptic membrane. In vitro, the above-mentioned enzymatic activities were completely suppressed by neuroactive agents such as catecholamines and phenothiazine derivatives. Examination using enzyme-histochemical techniques suggests that a single enzyme may be responsible for both above-mentioned enzymatic reactions. The functional significance of microglial cells in the normal central nervous tissue is discussed.

Notes: Summary The tight junctions along the medullary collecting duct in the kidneys of the rat and the rabbit were studied with freeze-fracture electron microscopy and quantitated according to the number of strands and the apico-basal depth (nm) of the junctions. The most elaborate tight junctions were found in the inner stripe of the outer medulla; rat: 10.6±0.8 strands and 205±24nm; rabbit: 11.6±2.4 strands and 291±55 nm. The elaboration of the tight junctions decreased continuously towards the papillary tip. Inner zone I; rat: 9.3±2.6 strands and 186±38nm, rabbit: 9.5±2.3 strands and 247±59nm. Inner zone II; rat: 7.1±2.2 strands and 129±32nm, rabbit: 8.5±1.4 strands and 199±26nm. Inner zone III; rat: 6.0±1.6 strands and 111 + 19 nm, rabbit: 7.0±1.5 strands and 183±43 nm. In the inner zone III comprising the papillary tip tight junctions with only 1–3 strands were not infrequently seen. Preliminary findings in the kidney of the golden hamster indicate a similar decline of junctional tightness along the collecting duct. These morphological observations suggest that the permeability of the paracellular pathway of the medullary collecting duct increases towards the tip of the papilla, especially in the rat. The functional implications for the medullary recycling of urea and electrolytes, and for the urinary concentrating mechanism are discussed. In addition, the tight junctions of the papillary epithelium are described.

Notes: Summary Immunoreactive luteinizing hormone-releasing hormone (LHRH)-like material has been demonstrated in the pineal gland of the adult rat. The objective of the present study was to examine the ontogenetic development of this LHRH-like substance in the rat pineal with the peroxidase-antiperoxidase (PAP) method of Sternberger. LHRH-like immunoreactive material was not observed in pineal glands of newborn rats. The amount of material increased progressively from the 6th–12th day of postnatal development. On day 12, the amount of LHRH-like immunoreactivity was consistent and comparable in all pineal glands of male and female animals examined.

Notes: Summary An ultrastructural and morphometric analysis was made of the nucleolar components in pinealocytes of 40 male Fischer rats sampled at eight times in an LD 12∶12 photoperiod cycle. Comparisons of results from the eight times showed variation in estimated mean volume of the granular component of ±29%, and of the fibrillar component ±11%, in relation to daily means. Peaks in mean volume of total nucleolus and its granular component occurred at 1 h of light. Near maximal and minimal mean volumes of the fibrillar component both occurred during both light and dark. Fibrillar centers (nucleolar organizer regions) of different sizes were found at all sampling times. It is concluded that temporal patterns in 24-h changes in the nucleolar components are most prominent in the granular component, and are more complex than suggested by changes in total nucleolar size or mean dimensions, and than represented by a simple biphasic circadian rhythm. Examples of different stages in the migration of the granular component, and of possible sites of nucleo-cytoplasmic transfer of nucleolar material, are described.

Notes: Summary Localization of hCG binding sites and the process of endocytosis in pseudopregnant rat ovaries were investigated by indirect electron-microscopic immunocytochemistry. Immature female rats were treated with pregnant-mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) to induce ovarian luteinization. Eight days after priming with PMSG-hCG and 1–6 h before sacrifice the animals were given another injection of hCG to bind the receptors. Receptor sites to hCG localized by reaction product were present in most luteal cells, but not in primary follicular cells. The receptor sites were distributed on luteal cell surfaces facing interstitial spaces. Endocytotic pits containing hCG binding sites were rarely seen 1 h after hCG injection. At 2 h, hCG and presumably its receptor were taken up within endocytotic vesicles with the evidence of reaction product coated on the vesicle wall. With time, fusion of endocytotic vesicles with lysosome occurred and the reaction product appeared in phagolysosomes. The reaction product was localized on phagolysosomal inner surface or in free granular form. These findings suggest that hCG and its receptors were internalized through endocytotic pits and endocytotic vesicles and delivered to lysosomes probably for degradation. An additional experiment for localization of acid phosphatase was also performed to delineate the lysosomes and phagolysosomes.

Notes: Summary The morphology and distribution of the sensory neurones of the pudendal nerve within the spinal ganglia of rats were investigated by use of horseradish peroxidase (HRP). The labelling was visualized in diaminobenzidine (DAB) or tetramethyl-benzidine (TMB)-stained sections. Injection of HRP directly into the pudendal nerve labelled perikarya predominantly in the sixth lumbar DRG (L6). Following injection of HRP into the scrotal skin, however, additional cells were labelled in L5 and SI. Labelling was invariably unilateral. Approximately equal numbers of small (〈30 μm) and large neurones (〉40 μm) were labelled following subcutaneous injections although injections into the nerve marked twice as many small cells as large cells. This suggests that, in the rat, most of the small-diameter fibres within the pudendal nerve ascend through L6. Although a cluster of neurones was observed in one experiment, the remaining 25 experiments did not reveal any somatotopic arrangement since the labelled perikarya were distributed evenly throughout the ganglion. Similar numbers of retrogradely labelled neurones (somatopetal transport of the tracer) were observed in both DAB- and TMB-stained sections, although TMB allowed the demonstration of anterograde (somatofugal) HRP transport by terminal labelling in the superficial laminae of the lumbar spinal cord, extending into laminae II–IV.

Notes: Summary The hypothalamo-extrahypophyseal neurophysin pathways (HEH) and the three hypothalamic nuclei secreting neurophysins, the supraoptic (SON), paraventricular (PVN) and suprachiasmatic (SCN) nuclei, of normal and hypophysectomized rats were studied by application of the immunoperoxidase procedure. Eight well-defined HEH pathways were recognized. Their main sites of projection were: lateral septum and subfornical organ (1 and 2); tractus diagonalis (3); medial nucleus of the amygdala and lateral ventricle (4); nucleus periventricularis thalami, nucleus habenulae lateralis and periaqueductal gray (5); periaqueductal gray, pineal organ, collicular recess and subependymal region of the fourth ventricle (6); dorsomedial nucleus and premammillary area (7); perimammillary region, corpus trapezoideum, ventral surface of medulla oblongata, nucleus tractus solitarii, nucleus commissuralis, substantia gelatinosa and formatio reticularis lateralis of the medulla oblongata and spinal cord (8). Neurophysin fibers of unknown origin were found in the frontal cerebral cortex. It was noted that in pathway 5 the amount of immunostainable material undergoes changes with age. The three neurophysin-secreting nuclei reacted differently following hypophysectomy. Among the HEH pathways the only one that seemed to be affected by hypophysectomy was that innervating the lateral septum. It is suggested that the neurons that survive hypophysectomy either do not project to the neural lobe or, alternatively, display axon collaterals projecting outside the neural lobe. Such a neuronal population could be the origin of the HEH pathways.

Notes: Summary The distribution of luteinizing hormone-releasing hormone (LHRH)-immunoreactive perikarya and processes was examined, in the untreated rat, with the unlabeled antibody enzyme method of immunocytochemistry on thick 50 μm vibratome sections. LHRH neurons were primarily observed in the preoptico-anterior hypothalamic and septal areas. Projections from these cell bodies to the median eminence form three distinct pathways, one laterally along the course of the optic tracts, one medially through the periventricular stratum of the third ventricle, and one through the tractus infundibularis. In addition, some of these cell bodies project to the organum vasculosum of the lamina terminalis (OVLT) and the subfornical organ (SFO). LHRH immunoreactive neurons were also noted in the anterior olfactory regions; they project along the medial olfactory tract to the olfactory bulb.

Notes: Summary Regeneration of splenic tissue after autologous subcutaneous implantation provides a useful model for studying the development of splenic tissue. The development of the various non-lymphoid cells of the white pulp in the rat is described. It appears that regeneration of the implants is initiated by ingrowing vessels and a newly formed reticulum, which forms the microenvironment for the homing lymphocytes. Marginal metallophils are found at their characteristic location at the inner border of the marginal sinus five weeks after implantation. Trapping of antigen-antibody complexes reappears when the first primary follicles can be recognized.

Notes: Summary Autoradiographic studies showed that in the rat hippocampus “microglia-like reactive cells” (MRC) and astrocytes are capable of proliferation in response to kainic acid (KA)-induced lesions. A marked increase in the number of labeled MRC was observed during the first four days after the induction of the KA-lesion. A proliferative response of astrocytes occurred at two days after the KA-lesion. After the induction of a KA-lesion brain macrophages and oligodendrocytes were only slightly labeled with 3H-thymidine. It appears likely that MRC is the main cellular element responding to this type of lesion.

Notes: Summary The ultrastructure of tight junctions between uterine luminal epithelial cells of the pregnant rat was studied by the freeze-fracture technique. On day 5 of pregnancy, the day of implantation, the region of tight junctions extended three times as far down the lateral cell membrane as on day 1 of pregnancy, and the strands of the complex interlinked more frequently. These observations suggest that tight junctions on day 5 may be more efficient in preserving the contents of the uterine lumen from dilution or escape than at earlier times of pregnancy.

Notes: Summary Cholecystokinin-like immunoreactivity has been demonstrated by radioimmunoassay and immunocytochemistry in the spinal cord of various mammals, in particular in nerve fibers of the superficial layers of the posterior column, but had not been detected in neuronal cell bodies. We report immunohistochemical evidence for the presence of a group of cholecystokinincontaining neuronal cell bodies in the lumbar spinal cord of the rat. This group of cells is only visualized after direct injection of colchicine into the spinal cord and is located near the central canal in the intermedio-medial nucleus of area X of Rexed.

Notes: Summary The morphology and localisation of IgM- and IgG-containing cells in the spleen of rats immunized with sheep red-blood cells (SRBC) were studied by combining immunohistochemical reactions with routine histological and histochemical methods. It was shown that IgM cells occur only in the outer periarteriolar lymphocyte sheath (PALS), whereas IgG cells are present throughout the whole PALS. It has been concluded that these cell types have different routes of migration. Both IgM- and IgG-containing cells were found in the germinal center in close relation with the extracellular immune complexes. The significance of this localisation is discussed.

Notes: Summary The morphological characteristics and distribution of the somata of serotonin-containing neurons in the brainstem of rats and cats were studied by use of the peroxidase-anti peroxidase (PAP) immunohistochemical method employing highly specific antibodies to serotonin. Antibodies were raised in rabbits against an antigen prepared by coupling serotonin to bovine thyroglobulin and using formaldehyde as the coupling reagent. The distribution pattern of serotonin neurons observed in the present material is essentially in agreement with that described by other investigators who used the Falck-Hillarp method. In addition, this immunohistochemical technique revealed serotonin-containing perikarya in the following regions: 1) the periaqueductal gray, especially lateral to the nucleus raphe dorsalis, 2) the nucleus interpeduncularis, 3) the nucleus parabrachialis ventralis and dorsalis, 4) the field of the lemniscus lateralis, and 5) the reticular formation of the pons and medulla oblongata. The described immunohistochemical procedure makes it possible to study central serotonin neurons in detail without pharmacological pretreatment. The wide distribution of serotonin neurons demonstrated in this study should be considered when interpreting experiments dealing with the serotonin system.

Notes: Summary In the preinfundibular portion of male rat hypothalamus, the arcuate-median eminence region was examined after staining with anti-porcine ACTH 1-39 serum. In several cases, anti-β-endorphin serum was also employed. Both sera stain the same cell bodies. The cell bodies of the immunoreactive neurons are scattered in the subependymal layer, arcuate nucleus and lateral tuberal region. Fibers originating from these cell bodies are distributed extensively throughout these regions. They make synaptic contacts on immunonegative fibers in the arcuate nuclei, or terminate directly in the perivascular space in the internal layer of the median eminence. The fibers, however, are very scarce in the external layer of the median eminence, and do not directly terminate on the neurovascular contact surface. These findings suggest that the ACTH/β-endorphin neuronal system may serve neuronally and humorally as an intrahypothalamic constituent of the hypothalamic pituitary regulation system.

Notes: Summary In the Ham-Wistar rat, a paraganglion was found within the vagus nerve at the site of branching of the recurrent laryngeal nerve. Due to its location the name “paraganglion supracardiale vagi” is suggested. Fluorescence microscopy of the paraganglionic cells displays an intense yellow-green fluorescence indicating the presence of biogenic amines. Ultrastructurally, chief cells containing dense-core vesicles form three kinds of synaptic contacts (afferent, efferent and reciprocal) with enlarged, mitochondria-rich nerve endings.

Notes: Summary Growing rats (4 weeks old) were kept for 3 weeks at 11° C and 24° C respectively. The cold-adapted animals showed a significantly higher oxygen consumption (64%). Volume density of subsarcolemmal and interfibrillar mitochondria as well as volume density of fat droplets were estimated in M. soleus and the diaphragm of both groups. In cold-adapted animals, the total volume of mitochondria was significantly increased by 24% in diaphragm and 37% in M. soleus. The volume of subsarcolemmal mitochondria was almost doubled in each muscle, but the volume of interfibrillar mitochondria did not change significantly. The surface of the inner mitochondrial membranes per unit volume of mitochondrion in M. soleus was significantly increased both in interfibrillar and subsarcolemmal mitochondria, whereas the surface of the outer mitochondrial membranes per unit volume of mitochondrion was increased only in the subsarcolemmal mitochondria. The volume of fat droplets in the diaphragm and M. soleus of cold adapted animals increased significantly by 62% and 150% respectively.

Notes: Summary The ultrastructural changes associated with the compensatory hypertrophy of the zona fasciculata cells on monoadrenalectomized rats were investigated by stereological techniques. It was found that these subcellular changes display a different pattern from those underlying the ACTH-induced adrenocortical cell growth in the intact rats. This result supports the view that compensatory adrenal hypertrophy does not involve activation of the hypothalamo-hypophyseal-adrenal axis.

Notes: Summary The localization of serotonin (5-HT)-immunoreactive nerve fibers in the cerebellum of the rat and cat was investigated by means of the peroxidase-anti-peroxidase (PAP) method using highly specific antibodies to 5-HT. Serotonin-containing nerve fibers were distributed throughout the entire cerebellum including the deep cerebellar nuclei, while 5-HT-positive neuronal somata were not detected in the cerebellum of either species. A different pattern of 5-HT innervation was found among the three layers of the cerebellar cortex. There were also interspecific differences in the pattern of distribution of 5-HT. In the rat, the pool of 5-HT nerve fibers mainly consisted of tangential elements, which were predominant in the molecular layer, while in the cat only a few 5-HT fibers were found in the molecular layer of the cerebellar cortex; dense networks of 5-HT nerve fibers were present in the granular layer. Some differences are evident in the pattern of distribution of 5-HT fibers in cerebellar regions classified on an anatomical and functional basis.

Notes: Summary Glomus cells from carotid bodies of adult rats dissociated by means of collagenase or collagenase + trypsin were used to study by electron microscopy the endocytotic uptake of cationized ferritin (CF) tracer into subcellular compartments. The glomus cells were incubated with the tracer (1) in a basic salt medium (BM), or (2) in the BM into which calcium ionophore A23187 had been added, or (3) in a potassium-rich medium. Incubation of the cells in BM containing CF for 30 min resulted in attachment of the tracer to the cell membrane and uptake of a few solitary tracer particles into small vesicles and multivesicular bodies. No uptake into the cisternae of the Golgi apparatus was observed. Further incubation in BM containing CF for another 30 min resulted in increased uptake of the tracer into small vesicles and multivesicular bodies. A similar pattern of uptake was observed when the dissociated glomus cells were first preincubated in BM with CF for 30 min and then incubated for 1 min or 30 min in the BM solution containing both the ionophore and CF. Upon such incubation, CF particles were seen to penetrate into coated pits and sites of exocytosis at the cell surface. When the 30-min preincubation in BM was followed by incubation in a CF-containing potassium-rich medium for 15–30 min, uptake into vesicles, small lysosomes and occasionally also into profiles of the smooth endoplasmic reticulum was seen. Endocytotic mechanisms of the glomus cells are outlined.