IDENTIFICATION OF A VESICULAR POOL OF CALCIUM CHANNELS IN THE BAG CELL NEURONS OF APLYSIA-CALIFORNICA

To study the molecular mechanism of calcium current modulation in the bag cell neurons of Aplysia californica, we have identified calcium ch annel subtypes expressed in these cells and analyzed their distributio n using channel-specific antibodies. Using PCR to amplify reverse-tran scribed RNA from bag cell clusters, we identified two classes of calci um channel alpha(1) subunit. One, BCCa-I, belongs to the ABE subfamily of calcium channels, whereas the other, BCCa-II, belongs to the SCD s ubfamily. Antibodies generated against the bag cell calcium channels r ecognize membrane proteins of similar to 210 and 280 kDa on immunoblot s. Both channels are expressed in the bag cell clusters as well as in other parts of the Aplysia nervous system. BCCa-II also localizes to g lia and muscle. The subcellular distribution of the two channel types is strikingly different. Anti-body staining of bag cell neurons in pri mary culture shows that BCCa-II is present on the plasma membrane, whe reas BCCa-I has a punctate, intracellular distribution consistent with a vesicular localization. The BCCa-I-containing vesicles are found in bag cell neuron somata and growth cones and occasionally in neuritic hotspots. Their distribution is similar but not identical to that of L ysoTracker Red, a marker for acidic organelles, but unlike that of den se-core vesicles containing egg-laying hormone. The vesicular channels may represent the protein kinase C-sensitive calcium channels of bag cell neurons that are believed to enhance hormonal release during elec trical activity.