Reducing ischaemia/reperfusion injury through δ-opioid-regulated intrinsic cardiac adrenergic cells: Adrenopeptidergic co-signalling

AimsThe purpose of this study was to determine whether intrinsic cardiac adrenergic (ICA) cells release calcitonin gene-related peptide (CGRP), exerting synergistic adrenopeptidergic cardioprotection.Methods and resultsIn situ hybridization coupled with immunostaining demonstrated that ICA cells exclusively expressed CGRP mRNA and co-expressed CGRP and δ-opioid receptor in human and rat left ventricular (LV) myocardium. Radioimmunoassay detected constitutive CGRP release from ICA cells in human and rat hearts. The δ-opioid agonist [D-Pen²⁵]-enkephalin (DPDPE) increased CGRP release from ICA cells in denervated rat heart. In an ischaemia/reperfusion rat model, pre-ischaemic treatment with DPDPE reduced infarct size (IS) by 51 ± 16 (P < 0.01). Co-infusion of β2-adrenergic receptor (β2-AR) and CGRP receptor (CGRP-R) antagonists increased IS by 62 ± 23 (P < 0.01) compared with saline and abolished DPDPE-initiated IS reduction. Pre-treatment of ICA cell-myocyte co-culture with the β2-AR/CGRP-R antagonists increased myocyte death rate by 24 ± 4 (P < 0.01) and abolished DPDPE-initiated myocyte protection against hypoxia/reoxygenation (re-O2). In the ICA cell-depleted myocyte culture, DPDPE did not confer myocyte protection. Supplementing ICA cell-depleted myocyte culture with β2-AR/CGRP-R agonists reduced hypoxia/re-O2-induced myocyte death by 24 ± 5 (P < 0.01), simulating endogenous neurohormonal effects of ICA cells. Western blot analysis showed that DPDPE markedly increased phosphorylated myocardial Akt levels. This effect was abolished in the presence of β2-AR/CGRP-R blockade. Terminal dUTP nick-end labelling staining analysis of the LV infarct zone demonstrated that DPDPE reduced myocyte apoptosis by 58 ± 19 (P < 0.05), an effect that was eliminated in the presence of β2-AR/CGRP-R blockade. Finally, echocardiography showed that DPDPE increased LV contractility in a manner dependent on β-AR/CGRP-R stimulation.ConclusionICA cells constitute a δ-opioid-regulated adrenopeptidergic paracrine system conferring robust cardioprotection through β2-AR/CGRP-R co-signalling, resulting in the activation of an anti-apoptotic pathway during ischaemia/reperfusion.