(Received for publication, March 20, 1996, and in revised form, June 3, 1996)

Karnam S. Murthy , David H. Coy and Gabriel M. Makhlouf

From the Departments of Physiology and Medicine, Medical College of Virginia, Richmond, Virginia 23298-0711 and the  Peptide Research Laboratory, Tulane University Medical Center, New Orleans, Louisiana 70112-2699

In COS-7 cells, all five cloned somatostatin receptors are coupled via inhibitory G proteins to activation of an unidentified phospholipase C- (PLC-) isozyme and inhibition of adenylyl cyclase. In the present study, intestinal smooth muscle cells (SMC) that express only one receptor type, sstr3, and possess a full complement of G proteins and PLC- isozymes were used to identify the PLC- isozyme and the G proteins coupled to it and to adenylyl cyclase.

Somatostatin-14 bound with high affinity to intestinal SMC; stimulated -myo-inositol-1,4,5-trisphosphate (IP₃) formation, Ca²⁺ release, and contraction; and inhibited forskolin-stimulated cAMP formation in a pertussis toxin-sensitive fashion. Somatostatin also stimulated phosphoinositide hydrolysis in plasma membranes. Only those somatostatin analogs that shared a high affinity for sstr3 receptors elicited muscle contraction. IP₃ formation, Ca²⁺ release, and contraction in permeabilized SMC and phosphoinositide hydrolysis in plasma membranes were inhibited (~80%) by pretreatment with antibodies to PLC-3 but not other PLC- isozymes, and by antibodies to G but not G. Inhibition of cAMP formation was partially blocked by antibody to G_i1 or G_o and additively blocked by a combination of both antibodies. Somatostatin-stimulated [³⁵S]GTPS-G complexes in plasma membranes were bound selectively by G_i1 and G_o antibodies.

We conclude that in smooth muscle sstr3 is coupled to G_i1 and G_o; the  subunits of both G proteins mediate inhibition of adenylyl cyclase, while the subunits mediate activation of PLC-3.

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