Orexin-A (OXA) is an excitatory hypothalamic neurotransmitter and ligand for Orexin Receptor-1 (OR1), isolated from a small group of hypothalamic neurons. OXA orchestrates different brain functions, and at the cognitive level some of the effects of insufficiency of OXA are well-known, for example in Parkinson’s disease. It is widely assumed that deteriorated cognitive processes are related to impaired network connectivity. However, little is known about the effects of OXA in network connectivity and synaptogenesis. Therefore, to obtain insight into this problem we designed experiments with two groups of networks of dissociated cortical neurons: one group incubated in a plain medium and another chronically treated with OXA. After one, two, three or four weeks in vitro we applied immunocytochemistry for detection of OXA, OR1 and synaptic marker synaptophysin. Shortly after plating, 91±8% of the neurons cultivated in a plain medium expressed OXA-immunoreactivity, which does normally not occur in vivo indicating that neurons may change their phenotype under non-natural culture conditions to develop synaptically coupled networks. The fraction of orexinergic neurons decreased to 33±21% after 4 weeks in vitro. OXA expression was highest in the first week of network formation, the period of maximum synaptogenesis, and then decreased and stabilized in the weeks thereafter. Our hypothesis that OXA plays a role in the network development as a synaptogenic factor was supported by higher levels, earlier onset, and sustained increase of synaptophysin-expression in experiments with chronic OXA application to the culture medium.
- Orexin A – Synaptophysin – Synaptogenesis – Dissociated cortical neuronal cultures – Immunocytochemistry – Rat