NEUROPEPTIDE DEGRADATION PRODUCES FUNCTIONAL INACTIVATION IN THE CRUSTACEAN NERVOUS-SYSTEM

The pentapeptide proctolin(in (Proct.; Arg-Tyr-Leu-Pro-Thr) is a modul atory transmitter found throughout the crustacean nervous system. No i nformation is available in this system, however, as to how the actions of this peptide are terminated. To study this issue in the crab Cance r borealis, we incubated exogenous proctolin (10(-5) M) with either th e thoracic ganglion (TG) or with conditioned saline (CS) that had been preincubated with the TG. We removed aliquots at standard time points for analysis by reverse-phase highperformance liquid chromatography ( HPLC). We found that over time the proctolin peak became progressively smaller, while three novel peaks appeared and increased in size. Comi gration experiments using HPLC indicated that the major novel peak was Proct. [2-5] (Tyr-Leu-Pro-Thr), while one of the two minor peaks was Proct. [3-5] (Leu-Pro-Thr). The other minor peak appeared to be Proct. [1-2] (Arg-Tyr), based on similar HPLC retention time to synthetic Pr oct. [1-2]. The reduction in the proctolin peak and the increase in th e Proct. [2-5] peak was prevented by co-incubation of proctolin with a ny one of several aminopeptidase inhibitors (10(-4) M) Proct. [1-2] an d Proct. [3-5] appeared to result from a diamino-peptidase-mediated cl eavage of proctolin. We tested whether N-terminal cleavage functionall y inactivated proctolin by coapplying proctolin (10(-8) M) and individ ual aminopeptidase inhibitors (10(-5) M) to the isolated stomatogastri c ganglion (STG). We found that these inhibitors significantly enhance d the proctolin excitation of the pyloric rhythm. Furthermore, applica tion of synthetic Proct. [2-5] to the STG had no effect unless high co ncentrations (>10(-6) M) were used, and neither Proct. [1-2] nor Proct . [3-5] (10(-4) M) influenced the pyloric rhythm. Our results indicate that proctolin is enzymatically degraded and thereby biologically ina ctivated in the crab nervous system, primarily by extracellularly loca ted aminopeptidase activity.