Identification and Functional Analysis of Crustacean Serotonin Receptors.

Constantly changing environments force animals to adapt by cycling through multiple physiological states. Plasticity in sensory, motor, and modulatory neural circuits is an essential part of these adaptive processes. Invertebrates with their accessible, identifiable neurons are excellent models for investigating the molecular and cellular mechanisms underlying state-dependent neural plasticity, and provide insight into similar processes in more complex systems. These properties have allowed highly detailed characterization of several crustacean circuits with respect to their connectivities, cellular properties, responses to various inputs, and outputs. Serotonin (5-HT) is an important neuromodulator in virtually every animal species. 5-HT signals are mediated primarily by a large family of metabotropic receptors on target cells that activate diverse intracellular signaling cascades. Although 5-HT’s effects on crustacean circuits have been studied in detail, the mediating receptors have been inaccessible until recently. Crustacean receptors had not been cloned and specific drugs for use in physiological experiments could therefore not be identified. Coupling properties of 5-HT receptor families are strongly conserved between phyla, but pharmacological profiles are not. The extent of pharmacological divergence among invertebrates is unclear, however, as no systematic functional profile of 5-HT receptors from related species has been determined. This work shows that orthologs of two 5-HT receptors, 5-HT2b and 5-HT1a, are highly conserved at the molecular, functional and pharmacological level between two distantly related decapod crustaceans, Panulirus interruptus and Procambarus clarkii. A suite of drugs was functionally characterized at Panulirus and Procambarus 5-HT2b and 5-HT1a receptors in cell culture, which were then used to investigate the roles of the receptors in pyloric cycle frequency modulation in the stomatogastric ganglion, a model central pattern generator. The two receptor subtypes were found to serve different roles in the circuit and their function depends on the initial state of the circuit. Finally, an antibody recognizing 5-HT1a was used to map the localization of this receptor within the crayfish nervous system. 5-HT1a is localized to somata and neuropil throughout the nerve cord, suggesting it may respond to synaptic, paracrine or neurohormonal 5-HT signals. The protein and mRNA expression levels are variable between individual animals, perhaps reflecting distinct physiological states