Neuroscience
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Immunofluorescence and immunogold labeling, together with sucrose gradient separation and Western blot analysis of microsomal subfractions, were employed in parallel to probe the endoplasmic reticulum in the cell body and dendrites of rat cerebellar Purkinje neurons. Two markers, previously investigated in non-nerve cells, the membrane protein p91 (calnexin) and the lumenal protein BiP, were found to be highly expressed and widely distributed to the various endoplasmic reticulum sections of Purkinje neurons, from the cell body to dendrites and dendritic spines. An antibody (denominated anti-rough-surfaced endoplasmic reticulum), which recognized two membrane proteins, p14 and p40, revealed a similar immunogold labeling pattern. ⋯ The latter receptor and the Ca2+ ATPase, known in other species to be concentrated in Purkinje neurons, exhibited bimodal distributions with a peak in the light and another in the heavy subfractions. A similar distribution was also observed with another lumenal protein, protein disulfide isomerase. Taken as a whole, the results that we have obtained suggest the existence in the endoplasmic reticulum of Purkinje neurons of two levels of organization; the first identified by widespread, probably general markers (BiP, p91, possibly p14 and others), the second by specialization markers, such as the inositol 1,4,5-triphosphate receptor and, possibly, p40, which appear restricted to areas where specific functions appear to be localized.