The objective is to study the chemosensory synapses of the carotid body (CB), formed by glomus cells and by the apposed terminals of the carotid body nerve (CBN), because: 1) Functionally, the glomus cells are probably presynaptic to the sensory nerve fibers, and their size (about 10 mu m) allows direct recordings from them; 2) Glomus cells contain and release ACh, dopamine and neuropeptides during sensory stimulation and one should analyze their membrane properties at rest and during secretory activity; 3) It is possible to study how released or applied transmitters affect the membrane of the postsynaptic CBN sensory terminals; and 4) There are marked trophic interactions between the glomus cells, the CBN sensory somata and their peripheral processes. Three lines of investigation will be pursued: (1) Voltage (whole cell or patch) clamping of isolated adult or cultured glomus cells to determine the nature of ionic channels involved in cellular activity. Intracellular ion-selective electrodes (liquid membrane type) should help reveal the cytoplasmatic ionic activities at rest and during receptor excitation or inhibition. (2) A study of the ionic channels responsible for the """"""""spontaneous depolarizing potentials"""""""" (SDPs) in CBN terminals to determine if they reflect postsynaptic effects of transmitters released from glomus cells. For this purpose we will use: a) small (1-2 mu m) fire polished electrodes, visually directed toward a nerve ending to suction part of its membrane into the electrode for whole terminal recordings; b) Artificial synapses between sensory neurons and glomus cells and made by transplanting or apposing CB fragments to the nodose ganglion to take advantage of electrotonic spread from nerve terminals. Glomus cells survive in the host, vagal fibers show chemosensitivity and functional connections between these elements are established; and c) We will record from neurons grown in co- culture close to glomus cells. (3) Trophic interactions between glomus cells and sensory fibers will be continued in a controlled environment by using cultured sensory ganglion neurons grown alone and with glomus cells since the neuronal electric and chemosensory properties in co-cultures differ from those recorded from neurons grown alone. An effort will be made to establish why the membrane of adult glomus cells is insensitive to (K+)o and why they do not generate action potentials although they originate from the neural crest. Cultured glomus cells may provide an answer since they become neuron-like, as do isolated cultured adrenal medullary cells. Thus, intracellular recordings and patch clamping will be made from freshly plated cells and after different times in culture to determine if there is a change in their ionic (K, Na and Ca) channels. These studies will be conducted with and without sensory neurons to determine possible effects of innervation.
