ROLE OF G-PROTEINS IN CILIARY PROCESS ADENYLYL-CYCLASE RESPONSES OF THE ALBINO RABBIT EYE

After intravitreal injections of cholera or pertussis toxin (CTX or PT X, 0.5-1 mu g/eye) into the albino rabbit eye, the in vitro responses of ciliary process adenylyl cyclase (AC) to isoproterenol, vasoactive intestinal peptide (VIP), and forskolin (FSK) were increased 21-40% fo r PTX, but for CTX-injected eyes AC responses to fluoroaluminate, VIP and FSK decreased 70-50%. The increased responses after PTX suggests t hat this toxin blocked an inhibitory G(i) control of AC that is presen t in the control tissue. However, prolonged (> 24 hr) in vivo exposure to CTX appears to down-regulate the AC enzyme. In contrast to the in vivo findings, AC responsiveness was unaffected by PTX pre-treatment o f membranes in vitro, while CTX pre-treatment increased basal activity (+600%), and the FSK response (+30%), but decreased responsiveness to fluoroaluminate, VIP and isoproterenol by 88-56%. Treatment of ciliar y process membranes with P-32-NAD and CTX or PTX followed by SDS-PAGE autoradiography of labelled proteins gave two bands for the G-protein alpha-subunits of Gs (45,56 kDa) and one broad band centered at 40 kDa for G(i)-type subunits respectively. Western blots using specific ant ibodies showed the presence of G(i) type I or III, but no detectable G (i) type II or G(o) in rabbit ciliary processes. We conclude that the changes in adenylyl cyclase enzyme responses after intraocular CTX or PTX may not correlate with cAMP levels and intraocular pressure effect s. However, the in vitro biochemical data on AC responses and on G-pro teins provide evidence for dual regulation of ciliary process AC by ac tivating and inhibitory G-proteins.