Vol. 286, Issue 2, 753-759, August 1998

Identification and Characterization of Muscarinic Receptors Potentiating the Stimulation of Adenylyl Cyclase Activity by Corticotropin-Releasing Hormone in Membranes of Rat Frontal Cortex

Pierluigi Onali and Maria C. Olianas

Section on Biochemical Pharmacology, Department of Neuroscience, University of Cagliari, Cagliari, Italy

In membranes of the rat frontal cortex, acetylcholine (ACh) and other cholinergic agonists were found to potentiate the stimulation of adenylyl cyclase activity elicited by corticotropin-releasing hormone (CRH). Oxotremorine-M, carbachol and methacholine were as effective as ACh, whereas oxotremorine and arecoline were much less effective. The facilitating effect of Ach was potently blocked by the M₁ antagonists R-trihexyphenidyl, telenzepine and pirenzepine and by the M₃ antagonists hexahydro-sila-difenidol and p-fluorohexahydro-sila-difenidol, whereas the M₂ and M₄ antagonists himbacine, methoctramine, AF-DX 116 and AQ-RA 741 were less potent. The mamba venom toxin MT-1, which binds with high affinity to M₁ receptors, was also a potent blocker. The pharmacological profile of the muscarinic potentiation of CRH receptor activity was markedly different from that displayed by the muscarinic inhibition of forskolin-stimulated adenylyl cyclase, which could be detected in the same membrane preparations. Moreover, the intracerebral injection of pertussis toxin impaired the muscarinic inhibition of cyclic AMP formation and reduced the Ach stimulation of [³⁵S]GTPS binding to membrane G proteins but failed to affect the facilitating effect on CRH receptor activity. The latter response was also insensitive to the phospholipase C inhibitor U-73122, the protein kinase inhibitor staurosporine and to the inhibitors of arachidonic acid metabolism indomethacin and nordihydroguaiaretic acid. These data demonstrate that in the rat frontal cortex, muscarinic receptors of the M₁ subtype potentiate CRH transmission by interacting with pertussis toxin-insensitive G proteins.