HIGH-DENSITY DISTRIBUTION OF ENDOPLASMIC-RETICULUM PROTEINS AND MITOCHONDRIA AT SPECIALIZED CA2+ RELEASE SITES IN OLIGODENDROCYTE PROCESSES

In oligodendrocyte processes, methacholine-evoked Ca2+ waves propagate via regions of specialized Ca2+ release kinetics (wave amplification sites) at which the amplitude and rate of rise of local Ca2+ signals a re markedly higher than in surrounding areas (Simpson, P. B., and Russ ell, J. T. (1996) J. Biol. Chem. 271, 33493-33501). In the present stu dy we have examined the effects of other phosphoinositide-coupled agon ists on Ca2+ in these cells, and the structural specializations underl ying regenerative wave amplification sites, Both bradykinin and norepi nephrine evoke Ca2+ waves, which initiate at the same loci and propaga te through the cell body and multiple processes via identical wave amp lification sites, Antibodies against type 2 inositol 1,4,5-trisphospha te receptors (InsP(3)R2) and calreticulin identify expression of these proteins in oligodendrocyte membranes in Western blots, Immunocytoche mistry followed by high resolution fluorescence microscopy revealed th at both InsP(3)R2 and calreticulin are expressed in high intensity pat ches along processes. Cross-correlation analysis of the profiles of lo cal Ca2+ release kinetics during a Ca2+ wave and immunofluorescence fo r these proteins along cellular processes showed that the domains of h igh endoplasmic reticulum protein expression correspond closely to wav e amplification sites, Staining cells with the mitochondrial dye, Mito Tracker(R), showed that mitochondria are only found in intimate associ ation with these sites possessing high density endoplasmic reticulum p roteins, and they remain in the same locations over relatively long pe riods of time. It appears, therefore, that multiple specializations ar e found at domains of elevated Ca2+ release in oligodendrocyte process es, including high levels of calreticulin, InsP(3)R2 Ca2+ release chan nels, and mitochondria.