AGGREGATION OF SODIUM-CHANNELS INDUCED BY A POSTNATALLY UP-REGULATED ISOFORM OF AGRIN

Agrin is involved in signaling the formation of high concentrations of acetylcholine receptors (AChRs) at the neuromuscular junction (NMJ), There are multiple isoforms of agrin attributable to alternative splic ing, and these isoforms are differentially expressed during developmen t and between tissues. The ability to cluster AChRs varies among the a grin isoforms. Sodium channels (NaChs) are also concentrated at the NM J. We have tested various agrin isoforms for their ability to induce f ormation of clusters of NaChs, We grew cocultures of dissociated adult rat muscle fibers with Chinese hamster ovary (CHO) cells that had bee n transfected with different isoforms of agrin. Using immunocytochemic al techniques, we determined that after 1 d in culture, CHO cells synt hesizing the neuronally expressed isoform with an eight amino acid ins ert (Agrin8) were able to form NaCh clusters at sites of contact betwe en the CHO cell and muscle cell, Clusters of NaChs could be formed any where along a muscle fiber, but more clusters were detected close to t he endplate where the endogenous level of NaChs was higher, None of th e other neuronal-specific agrin isoforms was able to cluster NaChs, Be cause Agrin8 is the only agrin isoform that is upregulated at birth wh en NaChs begin to cluster at the NMJ, we conclude that Agrin8 expressi on by motor neurons is a signal for NaCh clustering at the NMJ during normal development.