Cortical vesicle exocytosis in isolated cortices of sea urchin eggs: Description of a turbidometric assay and its utilization in studying effects of different media on discharge☆

摘要

Corticesofunfertilizedseaurchineggscanbeisolatedinsuspensionandwilldischargetheattachedcorticalvesicles(CVs)inresponsetocalcium.WedescribeasimpleturbidometricassayformonitoringtheCa2+-induceddischargeofthesevesiclesandalsocomparethedischargeofvesiclesisolatedinahighsaltmedium(primarilyKCl)withamediummorecloselysimulatingtheinternalmilieuofthecell(primarilypotassiumgluconateandglycine).Dischargeinresponsetocalciumissimilarinbothmedia,requiringapproximately6??Mcalciumforone-halfmaximaldischarge.Thereare,however,significantdifferencesinmorphologyandproteincompositionofthetwotypesofpreparations(moreproteinspresentintheglycinecortices)andalsointherateofdischargeofthevesiclesinresponsetocalcium(KClcorticeswitht126secasopposedto30secintheglycinecortices).TheglycinecorticesgraduallylosetheirabilitytorespondtocalciumbutretentionofcalciumsensitivityisconsiderablyaidedbyinclusionofATPinthemedia;ATPhasnoapparenteffectondischargeoftheKClcortices.Theglycinecortices,asopposedtotheKClcortices,exhibitedvariationincalciumsensitivityduringthebreedingseasonandinthenumberofvesicleswhichwouldnotbreakdowninresponsetoaddedcalcium(referredtoasrefractoryvesicles).Thequestionofwhichtypeofcortexpreparationmostcloselysimulatestheinvivosituationisdiscussed,andtheviewispresentedthattheglycinecorticesmostcloselyresembletheinvivosituation.