A long-term goal is to understand the consequences to innervated tissues (neuronal or non-neuronal) of neuronal dysfunction during development or after maturity, and to define both functional deficits and neuronal signals at the molecular level. The experiments outlined continue our program to delineate the role of the nervous system in regulating the functional mechanism and development of neurotransmitter and neuropeptide stimulated secretion in the rat parotid gland as a model system. The mechanism for the developmental lag in the onset of Beta-adrenergic-sensitive secretion and adenylate cyclase activation will be further investigated to relate the role of increases in catalytic subunit of adenylate cyclase to changes in Beta-adrenergic receptor number, Ns (guanine nucleotide regulatory subunit), and inhibitory coupling unit, Ni, using pertussis toxin. Tissue slices and dissociated cell or acini preparations will be used. Effects of neonatal sympathectomy and para-sympathectomy on regulation and functional maturation of this system will be studied. The properties and interactions of Alpha-adrenergic, muscarinic, substance P and VIP receptors and responses will be evaluated at the level of Ca++ independent activation of phosphatidyl inositide turnover, and the Ca++ dependent activation of 22Na+ influx, cGMP accumulation and exocytosis, in developing and mature animals and after sympathetic, parasympathetic or sensory denervation. The consequences of inactivating Ni in terms of receptor availability for activating other responses will be studied along with a possible role for calcium/phospholipid-dependent protein kinase, using phorbol ester in dissociated or cultured cells. Coordination of receptor development with its ability to activate a variety of responses will be explored along with the coordination and regulation of a single response through activation by a group of different receptors. The regulation of these processes by autonomic nerves will be studied. Cell culture systems for differentiated, dissociated cells and developing secretory cells will be set up to study the nature of factors regulating differentiation and supersensitivity. An understanding of these regulatory processes will help us in evaluating deficits due to specific neuronal loss or deficits in diseases such as familial dysautonomia, diabetic neuropathy, Parkinson's disease, amyotrophic lateral sclerosis, Alzheimer's disease and Huntington's disease.
