Acute Stimulation of Dissociated Cortical Neurons of Newborn Rats with Orexin A: Effect on the Network Activity

Abstract

Orexin A (OXA) and B are hypothalamic neu-ropeptides with recognized importance in the physiological regulation of various brain activities, including sleep/wakefulness, learning and memory, locomotion, auto-nomic control. Orexin activity is mediated by two types of receptors; OR1 binds OXA with higher affinity, while OR2 binds both ligands equally. There is a growing interest in OXA role in neurodegenerative diseases with respect to the non-motor symptoms such as sleep and attention disorders. Recent studies in Parkinson’s patients found 40% lower concentration of OXA in the frontal cortex and 25% reduction in the cere-brospinal fluid. Both the number of orexinergic neurons in the hypothalamus and the levels of orexin in the cerebrospinal fluid are reduced by 72% in the end-stage in mouse model of Huntington’s disease. Despite the extensive information about OXA expression and function in the nervous system of adults, data about the orexin system and its role in the functioning of developing neuronal networks are still insufficient. Neuronal cell cultures are often used as a model for brain inquiries; therefore, we undertook our study to investigate immunocyto-chemically the expression of OR1 in dissociated cortical neu-rons at various ages, and the acute effect of OXA application on the network activity of neurons, cultured on multi electrode arrays. Initially, control recordings were made after refreshing 50% (300 microliters) of the culture medium. After 2 hours, 300 microliters of medium were replaced again either with medium containing OXA (0.5-2 micromolar, n=8) or plain medium (n=5). Paired t-tests indicated a significant increase in the network activity after acute OXA application (p<0.003), but not after plain medium treatment (p>0.32), supporting the hypothesis that OXA stimulates neuronal activity in the cortex. These results indicate that potential drugs, based on OXA are attractive candidates for prevention and treatment of disor-ders associated with neuronal activity decline.