Pertussis toxin reverses prostaglandin E₂- and somatostatin-induced inhibition of rat parietal cell H⁺-production

In enzymatically dispersed enriched rat parietal cells we studied the effect of pertussis toxin on prostaglandin E₂ (PGE₂)- 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 PGE₂ (10^-9-10^-7 M) or somatostatin (10^-9-10^-6 M). PGE₂ inhibited histamine- and forskolin-stimulated [¹⁴C]aminopyrine accumulation but failed to alter the response to dbcAMP. Somatostatin was less effective and less potent than PGE₂ in inhibiting stimulation by histamine or forskolin and reduced the response to dbcAMP. Pertussis toxin completely reversed inhibition by both PGE₂ 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. PGE₂ 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.