Hindbrain lactostasis regulates hypothalamic AMPK activity and metabolic neurotransmitter mRNA and protein responses to hypoglycemia

Abstract

Nerve cell metabolic activity is monitored in multiple brain regions, including the hypothalamus and hindbrain dorsal vagal complex (DVC), but it is unclear if individual metabolosensory loci operate autonomously or interact to coordinate central nervous system (CNS) reactivity to energy imbalance. This research addressed the hypothesis that hypoglycemia-associated DVC lactoprivation stimulates hypothalamic AMPK activity and metabolic neurotransmitter expression. As DVC catecholaminergic neurons express biomarkers for metabolic monitoring, we investigated whether these cells are a source of lactate deficit signaling to the hypothalamus. Caudal fourth ventricle (CV4) infusion of the glucose metabolite l-lactate during insulin-induced hypoglycemia reversed changes in DVC A2 noradrenergic, arcuate neuropeptide Y (NPY) and pro-opiomelanocortin (POMC), and lateral hypothalamic orexin-A (ORX) neuronal AMPK activity, coincident with exacerbation of hypoglycemia. Hindbrain lactate repletion also blunted hypoglycemic upregulation of arcuate NPY mRNA and protein. This treatment did not alter hypoglycemic paraventricular oxytocin (OT) and lateral hypothalamic ORX mRNA profiles, but exacerbated or reversed adjustments in OT and ORX neuropeptide synthesis, respectively. CV4 delivery of the monocarboxylate transporter inhibitor, 4-CIN, increased A2 phosphoAMPK (pAMPK), elevated circulating glucose, and stimulated feeding, responses that were attenuated by 6-hydroxydopamine pretreatment. 4-CIN-infused rats exhibited increased (NPY, ORX neurons) or decreased (POMC neurons) pAMPK concurrent with hyperglycemia. These data show that hindbrain lactoprivic signaling regulates hypothalamic AMPK and key effector neurotransmitter responses to hypoglycemia. Evidence that A2 AMPK activity is lactate-dependent, and that DVC catecholamine cells are critical for lactoprivic control of glucose, feeding, and hypothalamic AMPK, implies A2 derivation of this metabolic regulatory stimulus.

Impact of recurrent hypoglycemic stress on hindbrain A2 nerve cell energy metabolism and catecholamine biosynthesis: modulation by estradiol

“…Precedent exposure to serial bouts of hypoglycemia caused opposite changes in basal A2 neuron DβH protein expression in estradiol-vs. oil-implanted OVX female rats, plus exacerbation of these divergent adjustments by renewed hypoglycemia. As these cells communicates hypoglycemia-associated cell energy imbalance signaling (Gujar et al 2014;Shrestha et al 2014), estradiol-dependent adaptive down-regulation of noradrenergic neurotransmission by A2 neurons implies that this hormone may promote a positive energy state during this recurring metabolic stress. Estradiol also regulates effects of recurrent hypoglycemia on enzyme proteins involved in energy metabolism and fatty acid synthesis.…”

Estradiol regulation of hypothalamic astrocyte adenosine 5′-monophosphate-activated protein kinase activity: Role of hindbrain catecholamine signaling

“…On days 14 and 16, animals were injected into the CV4 with the catecholamine neurotoxin 6-hydroxydopamine (6-OHDA) (N; 75 g/1.0 L/day; n = 10 E; n = 10 O) (Selvage et al, 2004) or vehicle (V; sterile apyrogenic water containing 0.2% ascorbic acid; n = 10 E; n = 10 O), as described (Gujar et al, 2014). This treatment paradigm results in significant loss of dorsal vagal complex catecholamine nerve cells without change in numbers of local neuropeptidergic neurons (Gujar et al, 2014).…”

“…This treatment paradigm results in significant loss of dorsal vagal complex catecholamine nerve cells without change in numbers of local neuropeptidergic neurons (Gujar et al, 2014). On day 18, one half of 6-OHDA-or vehicle-pretreated rats were injected sc with neutral protamine Hagedorn insulin (I; 12.5 U/kg bw), while the remainder were given sterile diluent (V) alone (Eli Lilly Co., Indianapolis, IN).…”

“…For each animal, three tissue sections were processed, per level, from rostral (−13.28 to −13.60 mm relative to bregma), commissural (−13.76 to −14.16 mm), and caudal (−14.36 to −14.86 mm) levels of the DVC for tyrosine hydroxylase (TH) immunoreactivity (-ir), as described (Gujar et al, 2014). Briefly, sections were washed with 0.05 M Tris-buffered saline (TBS, pH 7.6), pre-incubated (1 h) with 4.0% normal donkey serum (prod.…”

“…911724; Carl Zeiss MicroImaging), and immunolabeled for the astrocyte marker protein, glial fibrillary acid protein (GFAP), using described methods (Gujar et al, 2014). Briefly, tissues were fixed with acetone (10 min), blocked with 5.0% normal rabbit serum (1 h), and incubated (48 h; 4 • C) with goat polyclonal primary antibody (prod.…”