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Metabolic and serotonergic modulation of hypothalamic arcuate nucleus neurones in vitro

By Louise Saker


1. The effects of glucose on the electrophysiological properties of rat hypothalamic arcuate nucleus (ARC) neurones were investigated. Neurones were recorded in 10 mM (hyperglycaemic) and 2 mM (euglycaemic) glucose-containing aCSF. The major findings were that input resistance increased in 10 mM glucose, there was an increase in the activity of neurones in 2 mM glucose and there were a greater percentage of neurones expressing lh in 10 mM glucose. Subthreshold active conductances were differentially expressed in ARC neurones including: anomalous inward rectification Q. ), time- and voltage-dependent inward rectification 00, A-like transient outward rectification (IA) and T-type calcium-like conductance. Characterisation of the differential expression of these conductances may represent one way of functionally classifying ARC neurones. \ud 2. Whole-cell patch clamp recording techniques were used in isolated hypothalamic brain slice preparations to investigate the effects of 5-HT on ARC neurones. Bath application of 5-HT induced a membrane depolarisation in a sub-population of ARC neurones (30%), a response that persisted in the presence of TTX indicating a direct effect. 5-HT excited ARC neurones through three potential mechanisms: closure of one or more resting potassium conductances; activation of a non-selective cation channel, or a combination of the two; or activation of a pump in the membrane. This response was mediated through the 5-HT2A. 5-HT2B and/or 5-HT2C receptors revealed using a range of 5-HT receptor agonists and antagonists. 5-HT was shown to excite CART-expressing neurones suggesting an anorexigenic role for 5-HT, via 5-HT2 receptors at the level of the ARC.\ud 3.5-HT induced a membrane hyperpolarisation in a sub-population of ARC neurones (37%). The 5-HT-induced hyperpolarisation persisted in the presence of TTX indicating a direct effect on ARC neurones. 5-HT inhibited ARC neurones most likely through the activation of one or more potassium conductances,including an inwardly rectifying potassium conductance. Potential roles for 5-HTIA, 5-HTIB and 5-HT7 receptors were suggested from studies utilising 5-HT receptor agonists and antagonists. 5-HT inhibited orexigenic NPY/AgRP neurones, identified by their response to ghrelin and by their electrophysiological properties, suggesting an anorexigenic role for 5-HT, acting via 5-HTI and 5-HT7 receptors on NPY/AgRP neurones at the level of the ARC. \ud 4. The effects of feeding-related signals on hypothalamic neuropeptide expression were investigated using real-time-PCR. A new protocol measuring gene expression from hypothalamic explants was developed. Effects of GABA and AMPA on c-fos expression were investigated and subsequent studies showed leptin and glucose modulated the expression of NPY, POMC and AgRP, in fed and fasted animals. Further work is required to validate this novel approach to studying the central control of energy balance

Topics: RC0321, QM
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