AT2 receptors in cortical collecting ducts: a novel role in mediating ROMK-like K+ channel responses to high dietary K+?

angiotensin ii (ANG II) is perhaps one of the most extensively studied peptide “hormone” in cardiovascular, renal, and endocrine research and diseases. According to a recently published perspective from an analysis of the National Institute of Health Vascular Biology and Hypertension Branch-sponsored hypertension research by major experimental models used, almost half of total funding of research proposals somewhat directly or indirectly involves ANG II (4). Indeed, ANG II is like a bottle of well-brewed wine; the longer it is stored, the better it becomes. ANG II is widely recognized as an important endocrine, paracrine, and intracellular peptide that plays a key role in the regulation of cardiovascular and renal physiology and blood pressure homeostasis (5). In addition to its well-documented cardiovascular and renal hemodynamic actions, ANG II exerts powerful effects on the renal tubular transport of key electrolytes including Na+, bicarbonate, and K+, contributing to the maintenance of overall body Na+, K+, and fluid homeostasis (5). ANG II activates at least two major classes of G protein-coupled receptors (GPCRs), ANG II type 1 (AT1a and AT1b) receptors and ANG II type 2 (AT2) receptors, to elicit its biological or physiological effects. In the kidney as well as in other tissues, AT1 and AT2 receptors closely interact to play important counterbalancing “yin-yang” and/or complementary roles (2, 3, 12, 13). On the one hand, AT1 receptors act as the “yang” factor to mediate the effects of ANG II on vasoconstriction, aldosterone biosynthesis and release, glomerular filtration, and renal tubular Na+ and fluid reabsorption (5). On the other hand, AT2 receptors, as the “yin” factor, are simultaneously activated by ANG II to counteract renal hemodynamic and tubular responses to the activation of AT1 receptors by ANG II (1, 2). A fine balance between AT1 and AT2 receptor activation and their downstream signaling transduction in response to ANG II is critical to maintaining normal renal physiology, blood pressure, and body electrolyte and fluid balance. Over the last two decades, the roles of AT1 receptors and their signaling responses to ANG II have been extensively investigated in the kidney with enormous resources. This effort has significantly improved our current understanding of the physiological and pathological roles of ANG II via activation of AT1 receptors. In contrast, the roles of AT2 receptors and its downstream signaling responses to ANG II remain incompletely understood, in part due to a lack of enthusiasm and effort in the field as well as a general view that AT2 receptors may not be physiologically important. In a recent issue of the American Journal of Physiology Renal Physiology, Wei and colleagues (10) reported for the first time that ANG II acts on AT2 receptors to regulate renal outer medullary K+ (ROMK)-like channel activity in rat renal cortical collecting ducts (CCDs) in response to high dietary K+ intake. Specifically, these investigators showed that ANG II significantly increased ROMK channel activity in CCDs freshly isolated from rats fed a high-K+ diet but not a normal K+ diet. The effect of ANG II on ROMK channel activity was blocked by the AT2 receptor antagonist PD-123319 but not by the AT1 receptor blocker losartan and appears to involve the NO/cGMP/phosphodiesterase- and cAMP/PKA-dependent pathways (10). The results of this study are significant, because K+ homeostasis plays an important role in the regulation of various cellular and physiological functions. The reported findings provide new insights into a new physiological role for AT2 receptors in the kidney by regulating K+ homeostasis in response to altered dietary K+ intake. The purpose of this editorial commentary is not to discuss the very details of the overall results; instead, it only focuses on a number of key findings or issues as derived from this study for discussions. The first key finding of Wei and colleagues' study is that few AT2 receptors are expressed in CCDs of the adult rat kidney under a normal K+ diet, but AT2 receptor expression is substantially upregulated by a high-K+ diet. Currently, the expression and localization of AT2 receptors in the kidney remain unsettled issues in the ANG II/AT2 receptor research field and deserve further discussion. It is well recognized that the expression of AT2 receptors in human and rodent tissues is often developmentally related, with a high level of expression at the fetal and neonatal stages (3, 12). AT2 receptor expression generally disappears, although it may persist, in certain tissues of adult humans and rodents (3, 12). However, inconsistent results have been reported in the literature on AT2 receptor expression in kidney tubular segments, especially in CCDs of adult rodent kidneys (6–8, 10). Ozono et al. (7) previously showed that AT2 receptor expression was detected in the glomeruli, distal tubules, and interstitial cells of the adult rat kidney, which was upregulated in response to Na+ depletion. Reinhold et al. (8) did not detect AT2 receptor mRNA expression in freshly microdissected CCDs of the adult mouse kidney. In contrast, Miyata et al. (6) reported the expression of AT2 receptor mRNA and protein in both outer and inner medullary collecting ducts of the adult rat kidney. In the present study of Wei et al., the investigators used an alternative immunofluorescence labeling approach to clearly demonstrate AT2 receptor mRNA and protein in freshly isolated CCDs of the adult rat kidney, which increased three- to six-fold in response to 5% and 10% high-K+ diets, respectively. However, it should be pointed out that only 1 of 20 CCDs from normal K+ diet-fed rats was labeled by the anti-AT2 receptor antibody, suggesting very limited AT2 receptor expression occurred in CCDs of the adult rat kidney under physiological conditions. Although the mechanisms underlying the upregulation of AT2 receptors in CCDs by a high-K+ diet were not addressed by the study, the demonstration of AT2 receptor expression in CCDs does help explain the demonstrated effects of ANG II on ROMK channel activity via the activation of AT2 receptors in response to high K+ intake. The second key finding of the study of Wei et al. is that ANG II acts on AT2 receptors to stimulate ROMK-like channel activity in CCDs of the rat kidney in response to high-K+ diets, via the nitric oxide/cGMP pathway linked to the cAMP/PKA signaling pathway. Although the interaction between AT2 receptor activation by ANG II and nitric oxide/cGMP signaling is well recognized in the proximal tubule of the kidney (1), this represents a novel finding that has not been previously reported in CCDs of the rat kidney. In CCDs, ROMK channels are localized on the apical membranes of principal cells, which play an important role in the physiological regulation of renal K+ secretion (9). In contrast, K+ channels in basolateral membranes are involved in the regulation of cell membrane potentials and K+ recycling (9). In a previous study (11), the group showed that ANG II activated AT1 receptors to inhibit ROMK channel activity in split-open CCDs isolated from rats in response to low-K+ diet through PKC-, NADPH oxidase-, and protein tyrosine kinase-dependent signaling pathways. The recent study by Wei et al. demonstrates a new physiological response that clearly opposite to the above-mentioned effect of ANG II mediated by the activation of AT1 receptors in response to a low-K+ diet. These studies provide new evidence that AT1 and AT2 receptors may mediate opposite effects of ANG II on ROMK channel activity in CCDs of the kidney in response to altered dietary K+ intake. So what is the physiological relevance of the novel findings on the counterbalancing roles of AT1 and AT2 receptors in the renal regulation of K+ homeostasis? Specifically, it is not clear from the study of Wei et al. whether the changes in ROMK channel activity in CCDs in response to altered intrarenal or intratubular ANG II and dietary K+ intake would translate into changes in renal K+ secretion and therefore overall body and fluid K+ homeostasis. Wei and colleagues focused primarily on the roles of AT2 receptors and its downstream signaling transduction mechanisms in mediating the effects of ANG II on ROMK channel activity in response to high K+ intake at the CCD level. Clearly, the study was not designed to answer this important physiological question in an integrative physiological manner by determining the responses of systemic and intrarenal ANG II, aldosterone, plasma K+, and urinary K+ excretion, as well as other major electrolytes and blood pressure, to long-term altered dietary K+ intake. Additionally, the current dogma on the role of ANG II in mediating renal K+ homeostasis is that ANG II primarily acts on AT1 receptors to stimulate aldosterone biosynthesis and secretion in the adrenal glands, and, upon release, aldosterone acts in distal and collecting duct tubular segments to conserve Na+ and secrete K+. Since plasma K+ is a very sensitive and strong stimulator of aldosterone biosynthesis and secretion, it is expected that when dietary K+ intake is altered significantly, aldosterone biosynthesis and secretion from the adrenal glands will be impacted accordingly. The study of Wei et al. strongly suggests that ANG II may directly alter ROMK channel activity in CCDs via AT1 or AT2 receptors under physiological dietary K+ conditions. However, further studies may be necessary to determine the specific roles of AT1 or AT2 receptors in mediating the effects of ANG II on ROMK channel activity in CCDs of global and/or CCD-specific AT1a or AT2 receptor knockout mice chronically treated with either low- or high-K+ diets.