Immunocytochemical localization oftheplasmamembranecalcium pump,calbindin-D28k, andparvalbumin inPurkinje cells ofavian andmammalian cerebellum

Amonoclonal antibody produced against the humanerythrocyte plasma membrane calcium pump(PMCA) wasshowntoreact immunohistochemically with anepitope of thePMCAinavian andmammalian cerebellum. Western blot analysis ofpurified synaptosomes andhomogenates fromavian cerebellum revealed major immunoreactive proteins withmo- lecular masses (130kDaand138kDa)similar tothose of purified erythrocyte PMCA.Dual-imaging confocal immuno- fluorescence microscopy ofavian cerebellum showed that the PMCA antibody stained theperiphery ofthesomawhereas calbindin-D2sk waslocated inthecytosol. PMCA heavily stained themoredistal dendrites ofthePurki' ecells and, within theresolution ofthefluorescence procedure, colocalized withcalbindin-D%sk. Byusing alkaline phosphatase-conjugated second antibody, PMCAwasagain localized totheperipheral soma, toasegmental pattern indendrites, andtopresumed spiny elements. Thesomaperiphery anddendrites ofPurkiaje cells oftheratcerebellum werealso prominently stained with anti-PMCA antibody andcompared toparvalbumin localiza- tion. Dendritic depolarization anddendritic spiking behavior aresignificant Ca2+-dependent events ofPurki jecells. The rapid decline ofintracellular free Ca2+ after therapid rise time ofCa2+transients isconsidered tobeduetosequestration by Ca2+buffers, uptake byintracellular stores, andCae+ extru- sionmechanisms, thelatter afunction ofPMCAnowshown