Alteration of intracellular [Ca2+] in sea urchin sperm by the egg peptide speract. Evidence that increased intracellular Ca2+ is coupled to Na+ entry and increased intracellular pH.

Frontiers in electron probe microanalysis: application to cell physiology.

We describe a new cytochemical method for ultrastructural localization of intracellular calcium stores This method uses fluoride ions for in situ precipitation of intracellular calcium during fixation Comparisons made using oxalate, antimonate, or fluoride showed that fluoride was clearly superior for intracellular calcium localization in eggs of the sea urchin Strongylocentrotus purpuratus Whereas oxalate generally gave no intracellular precipitate and antimonate gave copious but random precipitate, three prominent calcium stores were detected using fluoride: the tubular endoplasmic reticulum, the cortical granules, and large, clear, acidic vesicles of unknown function The mitochondria of these eggs generally showed no detectable calcium deposits X-ray spectra confirmed the presence of calcium in the fluoride precipitates, although in some cases magnesium was also detected Rat skeletal muscle and sea urchin sperm were used to test the reliability of the fluoride method for calcium localization In rat skeletal muscle, most fluoride precipitate was confined to the sarcoplasmic reticulum Using sea urchin sperm, which transport calcium into the mitochondria after exposure to egg jelly to induce the acrosome reaction, the expected result was also obtained Before the acrosome reaction, sperm mitochondria contain no detectable calcium-containing precipitate Within 4 min after induction of the acrosome reaction, the expected result was also obtained Before the acrosome reaction, sperm mitochondria displayed many foci of calcium-containing precipitate The use of fluoride for intracellular calcium localization therefore appears to be a substantial improvement over previous cytochemical methods

Ultrastructural localization of intracellular calcium stores by a new cytochemical method.

http://www.jbc.org/content/259/22/13914.full.pdf

Measurement of plasma membrane and mitochondrial potentials in sea urchin sperm. Changes upon activation and induction of the acrosome reaction.

After ∼7–40 s following gamete fusion, a steadily increasing fraction of a sea urchin's zygotes initiate an activating calcium wave. The fertilization membrane then rises, the cell cycle resumes and development begins. This study focuses on the so-called latent period that occurs between the time that gamete fusion occurs and the initiation of the activating calcium wave. We inhibited calcium influx during this period by adding lanthanum or by reducing external calcium with a buffer at various time points after insemination. Both of these treatments blocked the activation of eggs that had not yet started a wave at the time of treatment. This indicates that an influx of calcium is needed during the latent period to induce egg activation. These results support the sperm conduit model of egg activation in the sea urchin, where calcium flows from the sea through the fused sperms' acrosomal process into a cortical region of the eggs' endoplasmic reticulum.

Role of calcium influx during the latent period in sea urchin fertilization

Energy-dispersive x-ray microanalysis was used to analyze changes in the subcellular distributions of Na, Mg, P, S, Cl, K, and Ca associated with the acrosome reaction of sea urchin sperm. Within 5 sec after induction of the acrosome reaction, nuclear Na and mitochondrial Ca increased and nuclear and mitochondrial K decreased. Uptake of mitochondrial P was detected after several minutes, and increases in nuclear Mg were detected only after 5-10 min of incubation following induction of the reaction. The results suggest that sudden permeability changes in the sperm plasma membrane are associated with the acrosome reaction, but that complete breakdown of membrane and cell function does not occur for several minutes.

D E Johnson

Papers

Changes in subcellular elemental distributions accompanying the acrosome reaction in sea urchin sperm.