DETACHED, PURIFIED NERVE-TERMINALS FROM SKATE ELECTRIC ORGAN FOR BIOCHEMICAL AND PHYSIOLOGICAL-STUDIES

Electric organs of skate (Raja species) dissociate to form populations of individual electrocytes when incubated in saline solutions contain ing collagenase, The rate of dissociation was highly temperature depen dent, with an apparent Q(10) of >6 in the range of 6 degrees-26 degree s C. The number of electrocytes per organ was relatively constant and independent of electric organ size, whereas mean cell diameters increa sed with organ size, The activities of two cholinergic marker enzymes, choline acetyltransferase (ChAT) and acetylcholinesterase (AChE), in extracts of whole fresh organs were much less than those reported for the electric ray Torpedo, suggesting a lower volume of terminals in th e organ. Electrocytes prepared from collagenase-treated organs had goo d resting potentials and generated postsynaptic evoked potentials. Spo ntaneous and electrode pressure-evoked miniature endplate potentials ( MEPPs) were readily recorded from isolated electrocytes. Incubation pe riods of more than 4 days in collagenase at 6 degrees C produced elect rocytes with good resting potentials and very low MEPP frequencies, in dicating denervation, Detachment of terminals and decreased MEPP frequ encies were concurrent, The time course of denervation was followed wi th the appearance of ChAT and AChE activities in a small particulate f raction derived from washed electrocytes. Peak activities of both enzy mes were seen at 4 days of incubation at 16 degrees C, but after 20 h at 16 degrees C, Electrocytes from 4-day, 6 degrees C incubations show ed detached, mitochondria-rich nerve terminals and dissociated Schwann cells. In unfixed preparations examined with Nomarski optics, isolate d nerve terminals were recognized and distinguished from nucleated Sch wann cells. Electron micrographs show that isolated terminals were sim ilar to attached terminals just before they dissociated. The MEPP freq uencies and evoked potentials were normal at terminals just before dis sociation. We conclude that the transmitter release process was normal in detached terminals and in terminals free of Schwann cells.