Investigating glucose responsive neuropeptide release using a static hypothalamic incubation system
S. Hussain1, E. Richardson2, G. Bewick1, S. Bloom1 & J. Gardiner1
The activity of certain hypothalamic neuronal populations is altered by changes in glucose. in-vitro studies have identified several glucose-sensitive neuronal populations in hypothalamic regions controlling energy homeostasis. Subsets of orexigenic arcuate nucleus Neuropeptide Y (NPY)-releasing neurones are known to be glucose responsive. These neurones may allow glucose to influence appetite. Cocaine and amphetamine regulated transcript (CART) is a neurotransmitter abundantly expressed in regions of the hypothalamus involved in energy homeostasis. At present, it is not clear if CART releasing neurones are glucose-sensitive. To assess hypothalamic neuropeptide release in response to changes in glucose, we investigated NPY and CART release by the hypothalamus at different concentrations of glucose, using a static hypothalamic incubation system.
Hypothalami from 20 male Wistar rats were incubated in artificial CSF (aCSF) for a 2-h equilibration period. The hypothalamic explants were then incubated for three 45 min periods in 600 μl aCSF containing 3, 8 (baseline) and 15 mM glucose in random order. Finally, the viability of the tissue was verified by 45-min incubation of aCSF containing 56 mM KCl. At the end of each incubation period, supernatants were removed and assayed for NPY and CART release by radioimmunoassay. Only explants that showed a greater secretion of NPY or CART with 56 mM KCl as compared to baseline were considered viable. Release of neuropeptide was expressed as percent of basal release.
There was a significant increase in NPY release with aCSF containing 3 mM glucose versus baseline glucose (3 mM: 285.2±92.8 vs 8 mM: 100±36.1 vs 15 mM 130.7±25.8). There was a decrease in CART release with aCSF containing 15 mM glucose versus baseline glucose (3 mM: 95.1±10.6 vs 8 mM: 100±6.8 and 15 mM: 78.8±3.9). These results suggest an important role for glucose-sensitive NPY-releasing hypothalamic neurones in mediating changes in energy homeostasis in response to glucose. They also suggest that subsets of CART-releasing hypothalamic neurones may be glucose responsive.
Declaration of interest: The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project.
Funding: This work was supported, however funding details unavailable.