The overall objective of this proposal is to elucidate the physiological, neurotransmitter and cellular mechanisms that operate in the rat carotid body in response to chemosensory stimuli, i.e., hypoxia, hypercapnia and acidity, using cultures of dissociated cells. The carotid body is intimately linked to the cardiovascular and pulmonary systems and speculated to be involved in certain cardiovascular disease states. The functional interactions between glomus (type 1) cells, the most numerous parenchymal cells of the carotid body, and their afferent nerve endings during chemosensory transduction are poorly understood due to: (i) difficulties in visualizing and obtaining reliable intracellular recordings from the small (10 um) glomus cells in situ, (ii) the complexity of the synaptic connections in vivo, (iii) the presence of multiple endogenous neurotransmitters and (iv) difficulty in separating effects due secondarily to circulatory changes. To simplify the circuitry, improve visibility, and obtain better control of the cellular and fluid environment, dissociated cell cultures of the rat carotid will be used. Further, this preparation is more favorable for obtaining reliable electrophysiological recordings from glomus cells by use of the patch-clamp technique. Isolated cells from the carotid body will be cultured alone or in combination with various sensory neurons, including chemosensory neurons from the petrosal ganglion. Experiments are designed to test for possible chemotransducer functions in glomus cells and will utilize electrophysiological, biochemical, light and electron microscopic procedures. Use will be made of the high resolution patch-clamp/whole-cell recording techniques to characterize the normal physiology and pharmacology of glomus cells and possible changes in ion-channel gating following exposure to chemosensory stimuli. The likely development of chemosensory contacts between petrosal sensory neurons and glomus cells will be followed in co-cultures by physiological and electron microscopic studies. Finally, the possible role of Ca2+ and H+ ions in chemosensory transduction and the involvement of various neurotransmitters will be investigated in these cultures.
