Activation of protein kinase C by serotonin: Biochemical evidence that it participates in the mechanisms underlying facilitation in Aplysia

1) Application of serotonin to Aplysia sensory neurons can result in facilitated synaptic transmission, both short- and long-term. This facilitation is likely to be produced by a complex set of molecular mechanisms: serotonin activates adenylate cyclase, increasing cAMP and protein kinase (Cedar and Schwartz, 1972); serotonin also changes the subcellular distribution of the Ca²⁺/calmodulin-dependent protein kinase (Saitoh and Schwartz, 1983). Recently, phorbol esters also have been shown to produce facilitation. We have therefore investigated how protein kinase C (PKC) participates in serotonin-mediated synaptic facilitation. 2) We found that the Aplysia genome encodes PKC, which is expressed in nervous tissue as at least two abundant transcripts (about 0.003% of the total message). Its inferred amino acid sequence is 85% homologous to that of enzymes from mammals and Drosophila, and over 95% homologous if compared to both. The specific activity of the Aplysia kinase is comparable to that found in rat brain, with similar reaction parameters and dependencies on phosphatidylserine (PS), Ca²⁺, diacylglycerol and phorbol esters. While PKC is found on neuronal membrane in the basal state, the PKC activators, Ca²⁺ and phorbol esters, further translocate the kinase to membrane in crude extracts of neuronal tissue. The amounts of membrane-bound PKC, as determined by ³H-phorbol-ester ginding, are greatest in neuropil and nerve. 3) Exposure of sensory neurons and their terminals in Aplysia pleural-pedal ganglia to facilitating doses of either phorbol ester or serotonin results in the translocation of PKC from cytosol to membrane, activating the enzyme. cAMP does not produce this translocation. Serotonin therefore must operate either by increasing Ca²⁺ or by stimulating the production of a lipid second messenger, possibly through an independent serotonin receptor. Persistent activation of PKC may result because the enzyme in association with membrane becomes autonomous, that is, independent of exogenous activators. 4) We have shown that the facilitatory transmitter serotonin produces changes in each of the three major second messenger pathways, and suggest that these intracellular signalling cascades, triggered by a single neurotransmitter should work toward a single behavioral end - enhancement of sensory neuron responses through increased excitability and facilitated release of transmitter.