The contractile state of Gastrointestinal (GI) smooth muscle and therefore GI motor function are under the constant control of both excitatory and inhibitory neurohumoral influences. To fully understand normal and disordered GI motor function, it is essential to elucidate the mechanisms by which excitatory and inhibitory influences are integrated to control smooth muscle contraction. Studies using isolate smooth muscle cells (SMCs) have demonstrated that single SMCs possess both excitatory (muscarinic) and inhibitory (Beta-adrenergic) receptors, and that simultaneous signals from these receptors can be integrated at the level of the single cell. Previous studies have explored Beta-adrenergic inhibition of SMC contraction. The studies proposed here will use isolated SMCs from the stomach of Bufo marinus (marine toad) to elucidate the muscarinic receptor-agonist interactions responsible for initiating smooth muscle contraction, the changes in intracellular [Ca2+] produced by muscarinic receptor activation and the physiologic end response (contraction) produced by muscarinic receptor activation. This approach is unique since all of these processes can be assayed in intact SMCs under identical conditions, making direct correlations possible. A number of techniques including radioligand binding assays, assays of intracellular [Ca2+] concentration with quin2 and assays of SMC contraction with the Coulter counter will be done. In addition, these studies propose to study how SMCs integrate excitatory (muscarinic) and inhibitory (Beta-adrenergic) signals to produce a contractile response. Since intracellular free [Ca2+] and cAMP appear to be biochemical mediators responsible for altering SMC contractile state, I will explore how muscarinic and Beta-adrenergic receptor activation may interact to modulate these signals. Of particular importance is how Beta-adrenergic receptor activation and; therefore, increases in cAMP may influence intracellular free [Ca2+] in resting and muscarinic stimulate SMCs. These studies are designed to explore issues of great physiologic and therapeutic importance; that is how neurohumoral agents modulate smooth muscle function. Alterations in these control mechanisms may be responsible for such illnesses as diffuse esophageal spasm or idiopathic intestinal pseudoobstruction. By taking this unique approach, new and useful information will be obtained.
