Ghrelin is a 28 amino acid peptide hormone secreted from X/A-like cells in the oxyntic mucosa of the stomach that has central actions to stimulate growth hormone release and orexigenesis [1]. The cognate receptor for ghrelin, GHSR-1a, is a 7-transmembrane G-protein coupled receptor that mediates most of the biological actions of ghrelin [2]. The principal physiological function of GHSR-1a was previously thought to be stimulation of growth hormone release from pituitary. However, GHSR-1a is expressed in many cell types including cardiomyocytes, myocardium, vascular endothelium, and monocytes raising the possibility that ghrelin may also have physiological actions in peripheral tissues. Indeed, ghrelin has direct peripheral actions (independent of growth hormone) to regulate cardiovascular homeostasis, inflammation, adipocyte function, bone formation, and gastric motility. The cardiovascular phenotypes of ghrelin null mice, ghrelin receptor null mice, and humans with ghrelin receptor mutations have not been reported to date. However, ghrelin has direct vasodilator, cardiotropic, cardioprotective, and anti-inflammatory actions [3].

      Intravenous infusion of ghrelin acutely lowers blood pressure in humans and rats. However, in rats, the hypotensive effects of ghrelin appear to be independent of vasodilator actions of nitric oxide (NO). Furthermore, studies in vivo [4] and ex vivo [5] suggest that ghrelin has acute NO-independent vasodilator actions. On the other hand, in patients with metabolic syndrome who have lower circulating ghrelin levels than healthy subjects, intra-arterial ghrelin infusion acutely improves endothelial dysfunction by increasing bioavailability of NO [6]. Moreover, a number of both central and ex vivo peripheral actions of ghrelin are NO-dependent. Therefore, mechanisms by which ghrelin exerts its peripheral and/or central hemodynamic actions have not been fully elucidated. In particular, the roles of vascular endothelium and NO in mediating cardiovascular actions of ghrelin remain unresolved. Intriguingly, ghrelin and insulin share some common actions to regulate energy homeostasis including stimulation of food intake and glucose metabolism [7]. The actions of insulin to regulate energy homeostasis are PI 3-kinase-dependent. Vasodilator actions of insulin to stimulate production of NO in vascular endothelium that help to couple metabolic and hemodynamic homeostasis are also PI 3-kinase-dependent [8]. Recently, it was found that ghrelin acutely activates endothelial nitric oxide synthase (eNOS) in vascular endothelium resulting in increased production of NO using PI 3-kinase-dependent signaling pathways shared in common with insulin [9].

Ghrelin-Stimulated Production of NO in Vascular Endothelial Cells Requires GHSR-1a and Involves Akt and e NOS, But Not MAP-Kinase

Treatment of aortic endothelial cells in primary culture with physiological concentrations of ghrelin increases NO production in a dose- and time-dependent manner that requires expression of GHSR-1a as well as activation of PI 3-kinase (but not MAP-kinase). Ghrelin-stimulated phosphorylation of Akt and eNOS at its Akt phosphorylation site also requires expression of GHSR-1a. Since PI 3-kinase-dependent phosphorylation of Akt and eNOS is known to increase production of NO, this strongly suggest that in vascular endothelial cells, ghrelin binds to its cognate receptor (GHSR-1a) resulting in activation of PI 3-kinase that then stimulates phosphorylation and activation of Akt that, in turn, phosphorylates and activates eNOS, resulting in increased production of NO. It is possible that ghrelin is also stimulating production of NO in endothelial cells through activation of other receptors known to bind ghrelin (e.g. CD36). However, this seems unlikely since reducing expression of GHSR-1a completely inhibits ghrelin-stimulated production of NO. These results are concordant with studies demonstrating that ghrelin stimulates phosphorylation of Akt in cardiomyocytes [10] and human endothelial cells [11]. Importantly, these data are fully consistent with human studies demonstrating NO-dependent vasodilator actions of ghrelin [6].

      The post-receptor signaling pathway used by ghrelin to stimulate production of NO in vascular endothelium is partially overlapping with that of insulin (insulin receptor/IRS-1/PI 3-kinase/PDK-1/Akt/eNOS) [8]. Thus, ghrelin joins the growing list of hormones involved with regulation of metabolism including insulin [12], estrogen [13], leptin [14], adiponectin [15], HDL [16], and DHEA [17] that acutely activate eNOS in vascular endothelium by a PI 3-kinase-dependent signaling mechanism leading to phosphorylation of eNOS by Akt. Metabolic and vasodilator actions of insulin are both regulated by highly parallel signaling pathways requiring PI 3-kinase (but not MAP-kinase). This represents one potential mechanism underlying the ability of vascular actions of insulin to help couple metabolic and hemodynamic homeostasis. Thus, ghrelin may also have functions to couple metabolic and cardiovascular physiology.

Ghrelin Stimulates Phosphorylation of MAP-Kinase but Not Secretion of ET-1 from Vascular Endothelial Cells

Ghrelin, like insulin, acutely stimulates phosphorylation of MAP-kinase in vascular endothelial cells [9,18,19]. Interestingly, by contrast with insulin, ghrelin does not stimulate secretion of the vasoconstrictor ET-1 from endothelial cells (a MAP kinase-dependent function of insulin). Thus, ghrelin mimics the vasodilator but not the vasoconstrictor actions of insulin in endothelial cells.

      Insulin resistance in diabetes and obesity is characterized by pathway-selective impairment in PI 3-kinase-dependent insulin signaling, but not MAP-kinase-dependent insulin signaling. Thus, in insulin-resistant states, compensatory hyperinsulinemia that serves to maintain euglycemia results in an imbalance between opposing PI 3-kinase-dependent vasodilator actions of insulin and opposing MAP-kinase-dependent vasoconstrictor actions of insulin that predisposes to endothelial dysfunction and hypertension. Ghrelin only mimics the PI 3-kinase-dependent vasodilator actions of insulin but not the opposing MAP-kinase-dependent vasoconstrictor actions of insulin. Therefore, ghrelin may have novel therapeutic potential for both metabolic and cardiovascular diseases characterized by reciprocal relationships between endothelial dysfunction and insulin resistance. Indeed, circulating ghrelin levels are abnormally low in insulin-resistant conditions including diabetes, obesity, metabolic syndrome, hypertension, coronary heart disease, and atherosclerosis. Moreover, polymorphisms in the human ghrelin gene are associated with diabetes, impaired glucose tolerance, and hypertension [20]. Interestingly, lifestyle interventions including exercise and/or therapeutic interventions that result in weight loss increase plasma ghrelin levels [21]. Thus, ghrelin-stimulated production of NO from vascular endothelium may contribute to the beneficial metabolic and cardiovascular outcomes resulting from the successful implementation of these strategies.

Conclusions

Novel vascular actions of ghrelin directly stimulate production of NO from vascular endothelial cells using PI 3-kinase-dependent signaling pathways that mimic those of insulin. These findings may explain, in part, molecular mechanisms underlying some beneficial cardiovascular actions of ghrelin. This raises the possibility that novel or existing therapeutic strategies to increase circulating ghrelin levels or ghrelin action may be beneficial for metabolic and cardiovascular diseases characterized by reciprocal relationships between insulin resistance and endothelial dysfunction.