Expression of vesicular glutamate transporter-2.1 in medaka terminal nerve gonadotrophin-releasing hormone neurones.

There are three paralogous genes for gonadotrophin-releasing hormone (GnRH) peptides of vertebrates in general. GnRH1, the protein product of gnrh1 gene, is the hypophysiotrophic neuropeptide, and is a critical regulator of gonadotrophin secretion, whereas GnRH2 and GnRH3 are regarded to have neuromodulatory functions. In some teleost species, the terminal nerve (TN) GnRH3 neuronal system, which expresses GnRH3, has been shown to project extensively throughout the brain and regulate the motivational state for some behavioural repertoires. In recent years, it has been considered that most, if not all, peptidergic and aminergic neurones synthesise and release more than one neurotransmitter, and the cotransmission of conventional small-molecule neurotransmitters, such as GABA, glutamate or acetylcholine together with neuropeptides, is regarded as a common feature of such neurones. For a functional characterisation of the GnRH3 neuronal system, we examined the possible co-expression of conventional neurotransmitters, GABA, acetylcholine and glutamate, in addition to GnRH in the TN-GnRH3 neurone by reverse transcriptase-polymerase chain reaction (RT-PCR) and in situ hybridisation of recently identified marker genes for neurotransmitters using a teleost fish medaka (Oryzias latipes). By RT-PCR and dual-label in situ hybridisation, we demonstrated the co-expression of GnRH3 and vesicular transporter for glutamate (VGluT) 2.1. in a single TN-GnRH3 neurone. We therefore suggest that the TN-GnRH3 neurones use glutamate as a cotransmitter of GnRH.