ALBERT

Notes: Summary A previous method of measuring the swelling pressure (ΔΠ g ) of the cytoplasmic gel of the giant axon ofLoligo vulgaris was refined. The estimates ofΔΠ g made with the improved method were consistent with those made with the earlier method. In these methods the activity of the solvent in the gel is measured by increasing the activity of the solvent in the internal phase of the gel by application of hydrostatic pressure to the gel directly. Comparable values for the activity of the solvent in the gel were obtained also by an alternate method, in which the deswelling of the gel is measured upon decreasing the activity of the solvent in the external phase by addition of a nonpenetrating high mol wt polymer (i.e., Ficoll). Additional support was obtained for the earlier suggestion thatΔΠ g contributes to the swelling and shrinkage pattern of the whole axon. In part, the new evidence involved two consecutivedirect measurements of intraxonal pressure. The first measurement was that of a mixed pressure composed ofΔΠ g andΔΠ m (ΔΠ m being the effective osmotic pressure due to the intra-extraxonal gradient in the activity of mobile solutes). The subsequent measurement was that ofΔΠ g alone. The latter measurement was made feasible by destroying the axolemma, thereby eliminating the contribution ofΔΠ m . An estimate ofΔΠ m was obtained by subtractingΔΠ g from the total pressure measured initially. TheΔΠ m determined by the above method was two orders of magnitude smaller than the theoretical osmotic pressure. This is consistent with theΔΠ m determined previously, where osmotic intra-extraxonal filtration coefficients were compared to the hydrostatic. The mixed pressure experiments lend credence to the idea that the substantial contribution ofΔΠ g to the water relations of the whole axon is due toΔΠ g being of the same order of magnitude asΔΠ m . The degree of free swelling of axoplasmic gels was studied as a function of pH, salt concentration, and hydration radius of the anion of the salt used. The swelling increased with an increase in the reciprocal of the hydration radius, a decrease in salt concentration, and at pH below or above ∼4.5. The nature of the constraints to the free swelling of axoplasm in axons immersed in seawater was studied. With the seawater employed, these constraints appeared to be due more to the retractive forces of the sheath than toΔΠ m .

Notes: Summary The hydrostatic (L p ) and osmotic (L PD ) filtration coefficients and the efflux rates of tritiated water were measured in the giant axon ofLoligo vulgaris. TheL p was 8 to 14×10−8 cm/sec/cm H2O and theL PD was two orders of magnitude smaller (3 to 6×10−10 cm/sec/cm H2O). In axons whose diameter was ∼500 μ, the time (t 1/2) required for a reduction in the axonal labeled water activity to one half its initial value was 38 to 48 sec. The rate limiting structure for solute flux was made ineffective by (1) storing the axon in isosmotic KF at 0–2 °C for one month to one year or by (2) fixing the axon in 2–4% glutaraldehyde for 3 to 7 hr. The criteria of ineffectiveness of the rate limiting structure for solute flux were (1) a reduction ofL PD to immeasurably low values, (2) the absence of electrical properties characteristic of a plasmalemma, and (3) a marked increase in the rate of efflux of Na22. In such impaired axons theL p and thet 1/2 of tritiated water efflux were unaffected. This independence of solute and solvent flux in conjunction with the finding that the hydraulic conductivity determined by bulk osmotic and hydrostatic pressure gradients is not equivalent (i.e.,L PD /L P ≪1) indicate that the rate limiting structures for solute and solvent flux are in series. Solvent fluxes appear to be surface-limited, not bulk-limited. We have been unable to resolve whether the surface structure involved in limiting solvent flux is the sheath (Schwann layer and adhering connective tissue) and/or the cortical layer of the axoplasmic gel.

Notes: Summary The volumetric elastic modulus of the sheath and the osmotic swelling pressure of the axoplasmic polymer network of the giant axon ofLoligo vulgaris were measured. Evidence was obtained that (1) the elastic modulus of the sheath, (2) the swelling pressure of axoplasm, and (3) theeffective osmotic pressure difference due to mobile solutes determine axonal volume. The contributions of the sheath and the axoplasm were significant because theeffective osmotic pressure due to mobile solutes was a small fraction of thetheoretical bulk osmotic pressure due to these solutes. The giant axon was converted from an imperfect to a near perfect osmometer by minimizing the contribution of the sheath and the axoplasmic gel.