Metabolism-independent sugar sensing in central orexin neurons - PubMed (original) (raw)

. 2008 Oct;57(10):2569-76.

doi: 10.2337/db08-0548. Epub 2008 Jun 30.

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Metabolism-independent sugar sensing in central orexin neurons

J Antonio González et al. Diabetes. 2008 Oct.

Abstract

Objective: Glucose sensing by specialized neurons of the hypothalamus is vital for normal energy balance. In many glucose-activated neurons, glucose metabolism is considered a critical step in glucose sensing, but whether glucose-inhibited neurons follow the same strategy is unclear. Orexin/hypocretin neurons of the lateral hypothalamus are widely projecting glucose-inhibited cells essential for normal cognitive arousal and feeding behavior. Here, we used different sugars, energy metabolites, and pharmacological tools to explore the glucose-sensing strategy of orexin cells.

Research design and methods: We carried out patch-clamp recordings of the electrical activity of individual orexin neurons unambiguously identified by transgenic expression of green fluorescent protein in mouse brain slices. RESULTS- We show that 1) 2-deoxyglucose, a nonmetabolizable glucose analog, mimics the effects of glucose; 2) increasing intracellular energy fuel production with lactate does not reproduce glucose responses; 3) orexin cell glucose sensing is unaffected by glucokinase inhibitors alloxan, d-glucosamine, and N-acetyl-d-glucosamine; and 4) orexin glucosensors detect mannose, d-glucose, and 2-deoxyglucose but not galactose, l-glucose, alpha-methyl-d-glucoside, or fructose.

Conclusions: Our new data suggest that behaviorally critical neurocircuits of the lateral hypothalamus contain glucose detectors that exhibit novel sugar selectivity and can operate independently of glucose metabolism.

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Figures

FIG. 1.

FIG. 1.

Effects of 2-deoxyglucose on the membrane potential of orexin neurons. A: A GFP-expressing orexin neuron identified in brain slice by epifluorescence (full description and validation of the GFP-labeling method is given by Burdakov et al. [11]) (top). Effects of 5 mmol/l bath-applied 2-deoxyglucose (2-DG) on membrane potential and resistance (resistance monitored as voltage deflections in response to hyperpolarizing current injections, see

research design and methods

) (bottom). B: Effect of 1 mmol/l bath-applied 2-deoxyglucose on membrane potential. C: Magnitudes of membrane potential hyperpolarization (Δ_V_m) induced by 1 and 5 mmol/l 2-deoxyglucose. ***P < 0.005, values are means ± SE, n = 4 cells for each condition.

FIG. 2.

FIG. 2.

Effects of 2-deoxyglucose on the membrane currents of orexin neurons. Whole-cell voltage-clamp recordings. A: Effect of 2-deoxyglucose on membrane current-voltage relationships. B: Current-voltage relationship of the net current activated by 2-deoxyglucose. The line is a fit of the GHK equation to the data (see

research design and methods

). Values are means ± SE, n = 4 cells.

FIG. 3.

FIG. 3.

Effects of 2-deoxyglucose on cortical neurons. A: Effect of 2-deoxyglucose on the membrane potential. Breaks in the trace correspond to where the recording was interrupted to expose the cell to voltage-clamp ramps (see

research design and methods

). Arrows show where the voltage-clamp ramps shown in B were taken. B: Effect of 2-deoxyglucose on membrane current-voltage relationship of the cell shown in A. No activation of K+ current is observed, but 2-deoxyglucose inhibits a current with a reversal potential of about −50 mV.

FIG. 4.

FIG. 4.

Effects of 2-deoxyglucose on orexin neurons in the presence of tolbutamide. A: Effect of 2-deoxyglucose on the membrane potential in the presence of intracellular (pipette) tolbutamide. Arrows show where the voltage-clamp ramps shown in B were recorded. B: Effect of 2-deoxyglucose on membrane current-voltage relationships of the cell shown in A.

FIG. 5.

FIG. 5.

Effects of lactate on orexin neurons. A: Effect of lactate on the membrane potential. Arrows show where voltage-clamp ramps shown in B were taken. B: Effect of lactate on the membrane current-voltage relationship of the cell shown in A. C: Current-voltage relationship of the net current inhibited by lactate.

FIG. 6.

FIG. 6.

Effects of glucokinase inhibitors and intracellular glucose on the glucose response of orexin neurons. A: Effect of 5 mmol/l glucose on the membrane potential in the presence of 10 mmol/l bath alloxan. B: Effect of 5 mmol/l glucose on the membrane potential in the presence of 10 mmol/l bath

d

-glucosamine. C: Effect of 5 mmol/l glucose on the membrane potential in the presence of 4 mmol/l bath _N_-acetyl-

d

-glucosamine (NAG). D: Cartoon showing experimental protocol (top). Effect on the membrane potential of 10 mmol/l interacellular glucose and 2.5 mmol/l extracellular glucose (bottom).

FIG. 7.

FIG. 7.

Effects of mannose on orexin neurons. A: Effect of 1 mmol/l mannose on the membrane potential (top). Effect of 4 mmol/l mannose on the membrane potential (bottom). Arrows show where membrane ramps shown in B were taken. B: Effect of mannose on membrane current-voltage relationships of the cell shown in A (bottom). C: Current-voltage relationship of the net current activated by mannose. The line is a fit of the GHK equation to the data (see

research design and methods

). Values are means ± SE, n = 6 cells.

FIG. 8.

FIG. 8.

Sugars that were not effective in hyperpolarizing orexin neurons. A: Lack of effect of 5 mmol/l

l

-glucose on the membrane potential and resistance, followed by normal response to 5 mmol/l glucose. B: Lack of effect of 5 mmol/l α-methyl-

d

-glucoside (α-MDG) on the membrane potential and resistance, followed by normal response to 5 mmol/l glucose. C: Lack of effect of 5 mmol/l fructose on the membrane potential and resistance, followed by normal response to 5 mmol/l glucose. D: Lack of effect of 5 mmol/l galactose on the membrane potential, followed by normal response to 5 mmol/l glucose.

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