QUANTITATIVE SINGLE-CELL REVERSE TRANSCRIPTION PCR DEMONSTRATES THAT A-CURRENT MAGNITUDE VARIES AS A LINEAR FUNCTION OF SHAL GENE-EXPRESSION IN IDENTIFIED STOMATOGASTRIC NEURONS

Different Shaker family alpha-subunit genes generate distinct voltage- dependent K+ currents when expressed in heterologous expression system s. Thus it generally is believed that diverse neuronal K+ current phen otypes arise, in part, from differences in Shaker family gene expressi on among neurons. It is difficult to evaluate the extent to which diff erential Shaker family gene expression contributes to endogenous K+ cu rrent diversity, because the specific Shaker family gene or genes resp onsible for a given K+ current are still unknown for nearly all adult neurons. In this paper we explore the role of differential Shaker fami ly gene expression in creating transient K+ current (I-A) diversity in the 14-neuron pyloric network of the spiny lobster, Panulirus interru ptus. We used two-electrode voltage clamp to characterize the somatic I-A in each of the six different cell types of the pyloric network. Th e size, voltage-dependent properties, and kinetic properties of the so matic I-A vary significantly among pyloric neurons such that the somat ic I-A is unique in each pyloric cell type. Comparing these currents w ith the I(A)s obtained from oocytes injected with Panulirus shaker and shal cRNA (lobster I-shaker and lobster I-shal, respectively) reveals that the pyloric cell I(A)s more closely resemble lobster I-shal than lobster I-shaker. Using a novel, quantitative single-cell-reverse tra nscription-PCR method to count the number of shal transcripts in indiv idual identified pyloric neurons, we found that the size of the somati c I-A varies linearly with the number of endogenous shal transcripts. These data suggest that the shal gene contributes substantially to the peak somatic I-A in all neurons of the pyloric network.