NEUROLIGANDS EVOKE CALCIUM SIGNALING IN CULTURED MYENTERIC NEURONS

Background. Myenteric neurons, which control gut motility and absorpti on, are heterogeneous in structure, electrophysiology, and neuropeptid e content. We hypothesized that myenteric neurons would display hetero geneous responses to neuroligands found in enteric tissue. We examined increases in intracellular calium ([Ca2+](i)) an important second mes senger, evoked by selected neuroligands in cultured myenteric neurons. Methods, Primary cultures of guinea pig myenteric neurons were charac terized by using a standard immunocytochemical stain for microtubule-a ssociated protein (MAP2). Increases in [Ca2+](i) were measured by digi tal imaging microscopy. Results expressed as mean +/- SEM, Student's t test. Results. Cultured myenteric neurons examined with phase contras t and Nomarsky optics extend processes and stain positively with the n euron-selective stain MAP2. Histamine did not evoke calcium mobilizati on in cult ured neurons, although progressive increases in [Ca2+](i) w ere seen with bradykinin, glutamate, serotonin, cholecystokinin, bombe sin, adenosine triphosphate (ATP) substance P, and acetylcholine. Ente ric neurons differed from one another in the ability to respond to one , two, or three of the agonist combinations tested. Time in culture di d not alter the percentage of neurons responding to ATP, substance P, or acetylcholine. ATP euoked equivalent [Ca2+](i) increases in neurons examined at the three time points, whereas significant decrements in neuronal calcium mobilization were seen after 8 days in culture with s ubstance P or acetylcholine (p < 0.05 versus 1 day in vitro). Conclusi ons. Cultured myenteric neurons extend processes and retain ex pressio n of neuron-specific antigens. Selected neuroligands found in enteric tissue increase [Ca2+](i) as a second messenger in subsets of myenteri c neurons. Heterogneity exists among cultured neurons in their ability to respond to ligand combinations. Changes in [Ca2+](i) induced by ne uroligands in myenteric neurons may be altered with time in vitro.