Over the years a number

Over the years, a number of studies have reported results concerning the behavior of the two endothelin receptor subtypes, ETA and ETB, that do not fit the classical model of two G protein-coupled receptors acting independently of one another. For example, in the rat anterior pituitary gland, both ETA and ETB receptors are expressed, but competitive binding studies indicated that ETB receptor-selective ligands competed for binding with endothelin-1 (ET-1) only when an ETA receptor-selective ligand was present. Simultaneous blockade of both receptor subtypes was necessary to inhibit clearance of ET-1 by astrocytes, with antagonists of individual receptor subtypes having no effect when administered alone. A similar cooperative interaction between the two receptors, necessitating blockade of both subtypes in order to abolish responses to ET-1, has also been noted in a variety of preparations involving vascular or airway smooth muscle. Systematic experiments by Just and colleagues demonstrated that the contributions of each receptor subtype to the renal vascular effects of ET-1 appear highly complex and are not merely additive. This may not be surprising given differences in receptor expression along the renal vascular tree combined with vasodilator and ET-1 clearance functions of endothelial-cell ETB receptors. However, those factors do not adequately explain the observation that either an ETA receptor-selective or an ETB receptor-selective antagonist was able to abolish afferent arteriolar constrictor responses to low concentrations of ET-1. In recent years, there has been much interest within the endothelin community regarding the possibility that ETA and ETB receptors may form homo- and heterodimers. Heterodimerization of endothelin receptors may provide an explanation for the results mentioned in the previous paragraph and might also account for other reports of atypical endothelin binding sites, although splice variants and post-translational modification of receptors should also be considered. It has been demonstrated with the aid of fluorescence resonance energy transfer analysis that, at least in transfected cells, ETA and ETB receptors can constitutively form heterodimers and homodimers. Furthermore, ETB receptors may internalize at a slower rate when present as ETA–ETB heterodimers. Consistent with this effect, HEK 293 Protease Inhibitor Cocktail (EDTA-Free, 200X in DMSO) transfected with both ETA and ETB receptors display a markedly prolonged (>4minutes) increase in [Ca] in response to exposure to ET-1 or the selective ETB receptor antagonist sarafotoxin 6c, compared with more transient (1–2minutes) responses of cells transfected with either receptor subtype alone. At least with endothelin receptors, most work to date has focused on heterodimerization of ETA and ETB receptors, rather than heterodimerization of endothelin receptors with receptors that bind other ligands. However, multiple combinations of receptor heterodimers have been described over the past 20 years, including dimers that bind the same ligand (for example, β-adrenoceptor–α-adrenoceptor dimers) or different ligands (for example, angiotensin AT–bradykinin B receptor dimers). Heterodimerization of receptors alters their ligand-binding properties and in some cases appears essential for activation of downstream signaling. Zeng and colleagues (this issue) now present results consistent with the possibility that endothelin ETB receptors and dopamine D receptors form heterodimers in renal proximal tubule cells from Wistar–Kyoto (WKY) rats. Dopamine receptors have previously been shown to heterodimerize with adenosine receptors in neurons, allowing adenosine agonists and antagonists to modulate the binding affinity and signaling activity of dopamine receptors. With both D and ETB receptors being expressed by the proximal tubule, it seems plausible that heterodimerization of ETB receptors with D receptors could also occur. Zeng and colleagues report that in a proximal tubular cell line derived from WKY rats, D receptors colocalize with ETB receptors in the plasma membrane. In contrast, this interaction appears to be disrupted in the corresponding cell line derived from spontaneously hypertensive rats (SHRs), with ETB receptors primarily localized in the cytoplasm and D receptors present in both the plasma membrane and cytoplasm. This difference may be functionally important, as the investigators reported that WKY rats produced a natriuretic response to intrarenal infusion of a selective D agonist, whereas SHRs, at least under conditions of high salt intake, did not. Moreover, the natriuretic effect of the D agonist in WKY rats could be attenuated by treatment with an ETB receptor antagonist, arguing in favor of some kind of functional cross-talk existing between ETB and D receptors in WKY rats. A question not addressed in the study is whether the relationship goes both ways—do D receptor antagonists impair the natriuretic effects of tubular ETB receptors? Also, what are the effects of agonists of D and ETB receptors on the actions mediated by one another?