Chronic opioids regulate K_ATP channel subunit Kir6.2 and carbonic anhydrase I and II expression in rat adrenal chromaffin cells via HIF-2α and protein kinase A

Abstract

At birth, asphyxial stressors such as hypoxia and hypercapnia are important physiological stimuli for adrenal catecholamine release that is critical for the proper transition to extrauterine life. We recently showed that chronic opioids blunt chemosensitivity of neonatal rat adrenomedullary chromaffin cells (AMCs) to hypoxia and hypercapnia. This blunting was attributable to increased ATP-sensitive K(+) (KATP) channel and decreased carbonic anhydrase (CA) I and II expression, respectively, and involved μ- and δ-opioid receptor signaling pathways. To address underlying molecular mechanisms, we first exposed an O2- and CO2-sensitive, immortalized rat chromaffin cell line (MAH cells) to combined μ {[d-Arg(2),Ly(4)]dermorphin-(1-4)-amide}- and δ ([d-Pen(2),5,P-Cl-Phe(4)]enkephalin)-opioid agonists (2 μM) for ∼7 days. Western blot and quantitative real-time PCR analysis revealed that chronic opioids increased KATP channel subunit Kir6.2 and decreased CAII expression; both effects were blocked by naloxone and were absent in hypoxia-inducible factor (HIF)-2α-deficient MAH cells. Chronic opioids also stimulated HIF-2α accumulation along a time course similar to Kir6.2. Chromatin immunoprecipitation assays on opioid-treated cells revealed the binding of HIF-2α to a hypoxia response element in the promoter region of the Kir6.2 gene. The opioid-induced regulation of Kir6.2 and CAII was dependent on protein kinase A, but not protein kinase C or calmodulin kinase, activity. Interestingly, a similar pattern of HIF-2α, Kir6.2, and CAII regulation (including downregulation of CAI) was replicated in chromaffin tissue obtained from rat pups born to dams exposed to morphine throughout gestation. Collectively, these data reveal novel mechanisms by which chronic opioids blunt asphyxial chemosensitivity in AMCs, thereby contributing to abnormal arousal responses in the offspring of opiate-addicted mothers.

Gene expression analyses reveal metabolic specifications in acute O2‐sensing chemoreceptor cells

“…; Salman et al . ), was practically the same in AM in comparison with CB or SCG cells (1.09‐fold change AM vs . SCG and 1.06‐fold change CB vs .…”

Localization of the delta opioid receptor and corticotropin-releasing factor in the amygdalar complex: role in anxiety

“…Previous studies using this DOPR antibody have demonstrated that DOPR immunoreactivity was abolished following preadsorption with antigenic peptide (Salman et al, 2013, Salman et al, 2014). In the present study to ensure antibody specificity, we performed control experiments and replicated these results with both immunohistochemistry and Western blotting technique.…”

Adrenergic Agents

“…Thus, under conditions of hypercapnia or hypoxia, the constrictor effect of catecholamines on brain vessels can disappear or decrease or a perverse (dilatatory) reaction occurs [8]. Under hypocapnia, noradrenaline causes cerebral vasoconstriction and decreased cerebral blood flow, apparently as a result of increased sensitivity of adrenergic vascular structures.…”