Our objective is to elucidate how mammalian cells perceive and respond to the signals in their environment. We have studied the effects of hormones and neurotransmitters which bind to and remain at extracellular receptors while transmitting information across the membrane. We have purified the beta-adrenergic receptor 2264-fold from C6 glioma cells, tagged the protein with a radioactive photoaffinity label, and demonstrated a single spot on autoradiographs of 2-D gels. Currently, we are purifying the receptor on a preparative scale to establish the amino acid sequence, clone the gene, and generate molecular probes to study regulation of receptor synthesis. Separate from receptors, we have found a new active transport system for a category of amines which includes beta-adrenergic antagonists such as propranolol and tricyclic antidepressants such as imipramine; transport is driven by a proton gradient across the plasma membrane maintained by a membrane-associated mgATPase. Transport by this system is fully active also in cells lacking beta-adrenergic receptors. Following the transportation and accumulation inside neurons and glial cells, tricyclic antidepressants such as imipramine can modulate the responses of both adenylate and guanylate cyclase cyclase to stimulation by neurotransmitters.
