[21] Application of antisense techniques to characterize neuronal ion channels in vitro

Publisher Summary Current gene cloning and genomic initiatives provide neuroscientists with an exponentially increasing bank of genetic sequence data that details the molecular identity of novel brain proteins. Often in the initial absence of selective ligands, the functional properties of the more recently cloned brain proteins remain unclear. The antisense approach conceptually offers a solution to this problem. Multiple mechanisms have been proposed that describe the molecular events that precipitate gene-specific knockdown by antisense oligonucleotides. These include inhibition of transcription, translational arrest, disruption of ribonucleic acid (RNA) processing, and RNase H–mediated transcript degradation. Despite the many questions that remain unanswered, antisense oligonucleotides have been applied successfully in the central nervous system (CNS) research. This chapter discusses their in vitro applications and describes both the strategies and methods that have been developed and applied successfully to the studies of neuronal voltage-dependent ion channels. Wherever possible, the effects of antisense treatment have been examined at multiple levels—that is, transcription transduction, messenger RNA (mRNA), protein and/or functionally by electrophysiological measurements. Each level of analysis affords complementary data that when taken together greatly facilitate the final interpretation of results.