Pertussis toxin reverses prostaglandin E 2- and somatostatin-induced inhibition of rat parietal cell H +-production

Abstract

In enzymatically dispersed enriched rat parietal cells we studied the effect of pertussis toxin on prostaglandin E 2 (PGE 2)- or somatostatin-induced inhibition of H +-production. Parietal cells were incubated in parallel in the absence (control cells) and presence of pertussis toxin (250 ng/ml; 4 h). [ 14C]Aminopyrine accumulation by both pertussis toxin-treated and control cells was used as an indirect measure of H +-production after stimulation with either histamine, forskolin or dibutyryl adenosine 3′,5′-cyclic monosphosphate (dbcAMP) alone and in the presence of PGE 2 (10 −9-10 −7 M) or somatostatin (10 −9-10 −6 M). PGE 2 inhibited histamine- and forskolin-stimulated [ 14C]aminopyrine accumulation but failed to alter the response to dbcAMP. Somatostatin was less effective and less potent than PGE 2 in inhibiting stimulation by histamine or forskolin and reduced the response to dbcAMP. Pertussis toxin completely reversed inhibition by both PGE 2 and somatostatin on histamine- and forskolin-stimulated H +-production but failed to affect inhibition by somatostatin of the response to dbcAMP. After incubation of crude control cell membranes with [ 32P]NAD +, pertussis toxin catalysed the incorporation of [ 32P]adenosine diphosphate (ADP)-ribose into a membrane protein of molecular weight of 41,000, the known molecular weight of the inhibitory subunmit of adenylate cyclase (G iα). Pertussis toxin treatment of parietal cells prior to the preparation of crude membranes almost completely prevented subsequent pertussis toxin-catalysed [ 32P]ADP ribosylation of the 41,000 molecular weight protein. It is concluded that in rat parietal cells pertussis toxin inactivates G iα by ADP-ribosylation thereby preventing inhibition mediated by the inhibitory subunit of adenylate cyclase. PGE 2 exerts its inhibitory effect on rat parietal cell function entirely by activating G iα. In contrast, somatostatin acts only in part via G iα, but also through additional pertussis toxin-insensitive mechanisms distal to or independent of the generation of cAMP.