ISSN:
0886-1544
Keywords:
flagellar dynein
;
cyclic nucleotides
;
sperm activation
;
Life and Medical Sciences
;
Cell & Developmental Biology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Medicine
Notes:
Methods of demembranation and reactivation of Lytechinus pictus sperm were developed that result in non-motile sperm which take on a stable bend of about 3.5 radians at the proximal end of the cell. The middle and distal portion of the flagellum is relatively straight or slightly bent in the same direction forming a somewhat “C” shaped sperm cell. In these studies, we refer to this characteristic shape as the quiescent form, and as opposed to “rigor wave” sperm, the quiescent form is induced and maintained by a relatively high concentration of MgATP2- (〉 0.2 mM). Other conditions important to the production and maintenance of the quiescent form in demembranated sperm include: starting with concentrated, undiluted sperm, maintaining low Ca++ in the demembranation buffer, using a minimum of 0.2 mM MgATP2- and pH of 7.9-8.1 in the reactivation buffer. Deviation from some of these conditions results in a dramatic increase in motile, asymmetrically beating sperm. Addition of 0.4 mM CaCl2 to the reactivation buffer increased the proximal bend angle to 5 radians. The induction and maintenance of the stationary bend is mediated by dynein activity: “rigor wave” sperm were transformed to the quiescent form upon 0.2 mM ATP addition; micromolar vanadate abolished the quiescent form by “relaxation” of the proximal bend; and the vanadate relaxed sperm were restored to quiescent form by catechol. Importantly, 20 μM cAMP activated motility of the otherwise quiescent-form sperm. Quiescent-form, demembranated sperm were also activated by mild trypsin digestion. These and other data suggest that the quiescent-form sperm are trapped at the end of the principal bend, and these data are consistent with the proposal that the single stationary bend results from asymmetry of active microtubule sliding [Gibbons and Gibbons, (1980): J. Cell Biol. 84:13-27].
Additional Material:
5 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/cm.970050304
