Elsevier

Brain Research

Volume 805, Issues 1ā€“2, 14 September 1998, Pages 41-54
Brain Research

Research report
Subgroups of hindbrain catecholamine neurons are selectively activated by 2-deoxy-d-glucose induced metabolic challenge

https://doi.org/10.1016/S0006-8993(98)00655-6Get rights and content

Abstract

Glucose is a major fuel for body energy metabolism and an essential metabolic fuel for the brain. Consequently, glucose deficit (glucoprivation) elicits a variety of physiological and behavioral responses crucial for survival. Previous work indicates an important role for brain catecholamine neurons in mediation of responses to glucoprivation. This experiment was conducted to identify the specific catecholamine neurons that are activated by glucoprivation. Activation of hindbrain catecholamine neurons by the antimetabolic glucose analogue, 2-deoxy-d-glucose (2DG; 50, 100, 200 or 400 mg/kg, s.c.) was evaluated using double label immunohistochemistry. Fos protein was used as the marker for neuronal activation and the enzymes tyrosine hydroxylase (TH) and phenethanolamine-N-methyl transferase (PNMT) were used as the markers for norepinephrine (NE) and epinephrine (E) neurons. 2-Deoxy-d-glucose (200 and 400 mg/kg) produced selective activation of distinct hindbrain catecholamine cell groups. In the ventrolateral medulla, doubly labeled neurons were concentrated in the area of A1/C1 and were predominantly adrenergic in phenotype. In the dorsal medulla, doubly labeled neurons were limited to C2 and C3 cell groups. In the pons, some A6 neurons were Fos-positive. Neurons in rostral C1, ventral C3, A2, A5 and A7 did not express Fos-ir in response to 2DG. Our results identify specific subpopulations of catecholamine neurons that are selectively activated by 2DG. Previously demonstrated connections of these subpopulations are consistent with their participation in the feeding and hyperglycemic response to glucoprivation. Finally, the predominant and seemingly preferential activation of epinephrine neurons suggests that they may play a unique role in the brain's response to glucose deficit.

Introduction

2-Deoxy-d-glucose (2DG) is a glucose analogue that competitively inhibits cellular glucose utilization [7]. Systemic or central administration of 2DG elicits a number of physiological responses that serve to redistribute and restore metabolic fuels 9, 23, 64. These responses to glucoprivation are crucial for survival because the brain is dependent on glucose as its metabolic fuel and on the continuous delivery of glucose by the blood.

Increased feeding [64]and adrenal medullary secretion 9, 23, 63are two responses to 2DG-induced glucoprivation. Increased adrenal medullary secretion is a neurally mediated response that facilitates delivery of glucose to the brain by stimulation of hepatic glycogenolysis and inhibition of insulin secretion 19, 71. Increased food intake replenishes deficient glucose reserves. Central norepinephrine (NE) and epinephrine (E) neurons appear to be involved in feeding and glycemic controls and may activate these controls during glucoprivation. Food intake and blood glucose concentrations are increased by central administration of NE and E 30, 45, 55, 66. Alpha-2 NE receptor binding in the paraventricular nucleus of the hypothalamus (PVH) is modulated by glucose concentration 10, 32. Blockade of alpha-noradrenergic receptors [40]or central administration of the selective catecholamine lesioning agent, 6-hydroxydopamine, significantly impairs glucoprivic feeding [68]. In addition, turnover of NE in hypothalamic terminals is increased by glucoprivation 3, 4, 47, 56and activity in these terminals is correlated with blood glucose across a range of glucose concentrations [65].

Despite evidence suggesting a role for NE neurons in glucoprivic feeding and adrenal medullary secretion, the particular catecholamine neurons involved in these responses have not been identified. Moreover, the relative importance of NE vs. E neurons in mediation of glucoprivic responses is not known since most pharmacological and neurotoxin manipulations that have been used to study the functions of NE and E neurons do not distinguish these two neurochemical phenotypes.

The c-fos proto-oncogene is now recognized as a reliable and sensitive indicator of neuronal activity induced by salient events in the internal and external environments [37]. Previously, we utilized immunohistochemical detection of Fos, the protein product of the c-fos gene, to map extended neuronal systems in brain, spinal cord and periphery that are activated by antimetabolic agents 48, 50. Results of mapping studies with 2DG [50]have shown that, among other sites of activation, 2DG induces Fos expression in the ventrolateral medulla, the nucleus of the solitary tract and the locus coeruleus, where NE and E cell groups are localized [24], as well as in the adrenal medulla 48, 53. However, these studies were not designed to determine whether 2DG induces Fos expression in the catecholamine neurons themselves. Therefore, in the present experiments, double label peroxidase immunohistochemistry was used to examine 2DG-induced Fos expression in neurons containing the catecholamine biosynthetic enzymes, tyrosine hydroxylase (TH) and phenethanolamine-N-methyl-transferase (PNMT). Since TH is present in both NE and E neurons, but PNMT is present only in E neurons, analysis of the co-distribution of Fos with each of these enzymes provides information regarding the involvement of both NE and E neurons in glucoprivic responses. Our results reveal anatomically localized, predominantly adrenergic subpopulations of neurons that are selectively activated by glucoprivation. These results support other findings indicating the existence in the hindbrain of anatomically distinct subgroups of catecholamine neurons that are distinguishable by their participation in different physiological functions 13, 61. Portions of this work have been reported previously in abstract form [52].

Section snippets

Materials and methods

Adult male Spragueā€“Dawley rats weighing 320ā€“340 g at the start of the experiment were obtained from Simonson Laboratories. Rats were housed individually in suspended wire mesh cages in a temperature-controlled room (21Ā±1Ā°C) illuminated between 0630 and 1830 h. Rats had ad libitum access to pelleted rat food and tap water, except during the 2-h drug test (see below). During the 1ā€“2 week period prior to the experiment, rats were extensively handled and habituated to the testing environment and

Results

Areas in which Fos- and TH-ir were quantitated are shown in Fig. 1. Results of quantitation of Fos-ir, TH-ir neurons and neurons doubly labeled for Fos- and TH-ir are shown for all areas in Table 1. Fos was not expressed in the areas under investigation in rats treated with saline control injections or with 50 mg/kg of 2DG. Therefore, these results are not included in Table 1. The threshold for expression of Fos in TH-ir neurons, and in the brain in general, was approximately 100 mg/kg. But as

Discussion

The present immunohistochemical results reveal that TH- and PNMT- containing neurons in caudal C1, C2 and dorsal C3 are strongly and selectively activated by glucoprivation. In C1 and C3, the neurons responsive to glucoprivation comprised distinct subpopulations distinguished by anatomical position within their respective cell groups. In C1, the 2DG-activated neurons were concentrated in the caudal 2/3 of the cell group. In C3, the activated neurons were those embedded in the floor of the

Acknowledgements

Supported by PHS # DK 40498 to SR.

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