Abstract
Glucokinase (GK), the hexokinase involved in glucose sensing in pancreatic β cells, is also expressed in hypothalamic tanycytes, which cover the ventricular walls of the basal hypothalamus and are implicated in an indirect control of neuronal activity by glucose. Previously, we demonstrated that GK was preferentially localized in tanycyte nuclei in euglycemic rats, which has been reported in hepatocytes and is suggestive of the presence of the GK regulatory protein, GKRP. In the present study, GK intracellular localization in hypothalamic and hepatic tissues of the same rats under several glycemic conditions was compared using confocal microscopy and Western blot analysis. In the hypothalamus, increased GK nuclear localization was observed in hyperglycemic conditions; however, it was primarily localized in the cytoplasm in hepatic tissue under the same conditions. Both GK and GKRP were next cloned from primary cultures of tanycytes. Expression of GK by Escherichia coli revealed a functional cooperative protein with a S0.5 of 10 mM. GKRP, expressed in Saccharomyces cerevisiae, inhibited GK activity in vitro with a Ki 0.2 µM. We also demonstrated increased nuclear reactivity of both GK and GKRP in response to high glucose concentrations in tanycyte cultures. These data were confirmed using Western blot analysis of nuclear extracts. Results indicate that GK undergoes short-term regulation by nuclear compartmentalization. Thus, in tanycytes, GK can act as a molecular switch to arrest cellular responses to increased glucose.
Highlights
Specific brain regions, such as the hypothalamus and brain stem, are able to detect and respond to changes in glucose concentration, triggering neuroendocrine responses that regulate feeding behavior [1]
There are four types of tanycytes: a1, a2, b1 and b2 [8]. a2 and b1 tanycytes are localized in the lower lateral wall of the third ventricle; they have extended cell processes that contact the neurons in the arcuate nucleus (AN), in particular neuropeptide Y (NPY) neurons [5,9,10] and pro-opiomelanocortin (POMC) neurons [11]. b2 tanycytes form the cerebrospinal fluid (CSF)-median eminence (ME) barrier, and their extended processes contact vessels devoid of the blood–brain barrier and are sometimes in direct contact with microvessels present in the ME [12]
Intracellular localization of GK in the hypothalamus and liver in response to glycemia In order to establish the levels of cerebral glucose reached in the hyperglycemic condition, glycorrhachia was measured in third ventricle canulated rats
Summary
Specific brain regions, such as the hypothalamus and brain stem, are able to detect and respond to changes in glucose concentration, triggering neuroendocrine responses that regulate feeding behavior [1]. Independent study in brain slices has shown that selective stimulation of tanycyte cell bodies by glucose and non-metabolizable analogs of glucose evokes robust ATP-mediated Ca2+ responses [15]. Both studies showed that Ca2+ waves were dependent on ATP release and subsequent activation of P2Y1 receptors. ATP or lactate release by tanycytes may modulate neuronal activity in hypothalamic areas associated with feeding behavior, including the arcuate nucleus (AN) and VMN, which are in close contact with these cells.
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