Immunohistochemical detection of α1E voltage-gated Ca2+ channel isoforms in cerebellum, INS-1 cells, and neuroendocrine cells of the digestive system

Polyclonal antibodies were raised against a common and a specific epitope present only in longer alpha 1E isoforms of voltage-gated Ca2+ channels, yielding an "anti-E-com" and an "anti-E-spec" serum, respectively. The specificity of both sera was established by immunocytochemistry and immunoblotting using stably transfected HEK-293 cells or membrane proteins derived from them. Cells from the insulinoma cell line INS-1, tissue sections from cerebellum, and representative regions of gastrointestinal tract were stained immunocytochemically. INS-1 cells expressed an alpha 1E splice variant with a longer carboxy terminus, the so-called alpha 1Ee isoform. Similarily, in rat cerebellum, which was used as a reference system, the anti-E-spec serum stained somata and dendrites of Purkinje cells. Only faint staining was seen throughout the cerebellar granule cell layer. After prolonged incubation times, neurons of the molecular layer were stained by anti-E-com, suggesting that a shorter alpha 1E isoform is expressed at a lower protein density. In human gastrointestinal tract, endocrine cells of the antral mucosa (stomach), small and large intestine, and islets of Langerhans were stained by the anti-E-spec serum. In addition, staining by the anti-E-spec serum was observed in Paneth cells and in the smooth muscle cell layer of the lamina muscularis mucosae. We conclude that the longer alpha 1Ee isoform is expressed in neuroendocrine cells of the digestive system and that, in pancreas, alpha 1Ee expression is restricted to the neuroendocrine part, the islets of Langerhans. alpha 1E therefore appears to be a common voltage-gated Ca2+ channel linked to neuroendocrine and related systems of the body.