Contractions are reported to be mediated at least partly by α_1A-adrenoceptors in a number of tissues including rat mesenteric artery (see Fig. 4.2), rat renal artery (also α_1D) [141], rat-tail artery [76, 140] and rabbit ear artery [45]. See also the following section on α_1D-adrenoceptors.

α₁-Adrenoceptors in blood vessels were subdivided based on their affinities for prazosin, WB 4101 and HV 723 into α_1H and α_1L [100]. α_1H-Adrenoceptors had high affinity for prazosin, and appeared to match the α_1A, α_1B, α_1D classification [101], whereas α_1L had low affinity for prazosin and did not seem to match the current molecular-cloning-based classifications. Under this classification, and often based on the low potency of prazosin, rabbit aorta, mesenteric and carotid arteries [100], guinea pig aorta, [101], rabbit cutaneous resistance arteries [123] and rat small mesenteric artery [137] were reported to contain α_1L-adrenoceptors.

Knockout (KO) of the α_1A-adrenoceptor abolishes α_1L-adrenoceptor pharmacology. The α_1A-adrenoceptor, expressed in the Chinese hamster ovary CHO-K1 cell line, displayed binding properties of the α_1A-adrenoceptor, but functionally, in terms of inositol phosphate accumulation, the receptor displayed properties of the α_1L-adrenoceptor [49]. Hence, the same receptor showed characteristics of both α_1A– and α_1L-adrenoceptors: in ligand-binding studies α_1A, but in functional studies α_1L. However, receptors reported to be α_1L-adrenoceptors can be demonstrated under slightly altered conditions to be α_1A-adrenoceptors. Hence, in the view of this author, it is likely that the α_1L-adrenoceptor is simply the native α_1A-adrenoceptor, at which prazosin shows low potency functionally [37] .

The α_1B-adrenoceptor has been somewhat a mystery receptor in vascular studies. Contractions are reported to be mediated at least partly by α_1Β-adrenoceptors in a number of tissues: rabbit corpus cavernosum [104] and rabbit cutaneous resistance arteries [123]. In studies of blood vessels from adrenoceptor KO mice, it was found that the major subtypes involved in contractions are α_1A and α_1D, but that the α_1Β-adrenoceptor plays a minor but clearly defined role in contractions in aorta, carotid, mesenteric and tail arteries [28]. For instance, contractions in rat-tail artery develop more slowly in α_1Β-adrenoceptor KO mice [28], so that subtle differences can be revealed following receptor KO. In the mouse mesenteric vasculature, α_1B-adrenoceptor KO resulted in diminished nerve but not agonist-induced vasoconstriction at 10 Hz but not 2 or 4 Hz [136].

In rat aorta, both α_1A– and a_1B-adrenoceptors are involved in trophic effects [149], and in mouse femoral arteries, employing gene knockout technology, it has been demonstrated that both α_1A– and α_1B-adrenoceptors are required for neo-intimal formation (no single knockout was effective) [68].

BMY 7378 is a selective antagonist at α_1D-adrenoceptors [58] and has been useful in identifying responses mediated by α_1D-adrenoceptors. Contractions are reported to be mediated at least partly by α_1D-adrenoceptors in a number of tissues including: rat aorta [1, 69, 110], rat iliac artery [110], rat mesenteric artery and pulmonary artery [69], rat renal artery (also α_1A) [141], rat carotid artery, mesenteric artery, aorta [140] and rabbit aorta (also possibly α_1A) [45]. In contrast, some studies of rat mesenteric artery did not report α_1D-adrenoceptor involvement in contractions [110]. The α_1D-adrenoceptor is reported to be constitutively active in the rat aorta, meaning that a response can occur in the absence of an agonist, and antagonists such as prazosin behave not simply as competitive antagonists but also as inverse agonists to inhibit that constitutive activity [6, 57].

Clearly, contractions in a number of tissues are mediated by more than one subtype of the α₁-adrenoceptor, and currently available subtype selective antagonists are often not selective enough to tease out clearly which receptors are present. In the perfused rat kidney, both α_1A– and α_1D-adrenoceptors mediate vasoconstriction [5], and in the external carotid circulation, the use of selective antagonists suggests that responses involve α_1A, α_1D, α_2A and possibly α_2C [143a]. Unlike the rat aorta, where the predominant receptor is α_1D, contractions of the mouse aorta clearly involve α_1A– in addition to α_1D-adrenoceptors [146]. Receptor KO studies in mice can help. In α_1D-adrenoceptor KO mice, there was a reduced vasoconstrictor response to both NA and nerve stimulation in femoral arteries [148], demonstrating the role of this receptor in neurotransmission (see later). In mouse mesenteric arteries, there is evidence from antagonist studies for the involvement of both α_1A– and α_1D-adrenoceptors in contractions [146], although the predominant α₁-adrenoceptor is the α_1A-adrenoceptor.

The role of the α_1A– and α_1D-adrenoceptor in contractile responses in a number of blood vessels may best be illustrated by Fig. 4.2. Figure 4.2 shows concentration–response curves for contractions produced by the α₁-adrenoceptor agonist phenylephrine in small mesenteric artery from wild-type (WT) and α_1D-adrenoceptor KO mice. KO of the α_1D-adrenoceptor changes the response from a relatively shallow concentration–response curve to a steep curve. The α_1D-adrenoceptor-mediated response is the component found in WT but not in α_1D-adrenoceptor KO mice. Note that low concentrations of phenylephrine (and indeed noradrenaline) selectively activate α_1D-adrenoceptors (Fig. 4.2). The high-affinity receptor is an α_1D-adrenoceptor. This is also true for noradrenaline: Noradrenaline has high potency at the α_1D-adrenoceptor and lower potency at the α_1A-adrenoceptor, so that knockout of the α_1D-adrenoceptor removes the high-potency component of the response to noradrenaline . The high affinity of this receptor for noradrenaline makes it the most likely candidate for the receptor in the neuroeffector region involved in the control of neurotransmission, and indeed blood pressure is reduced by deletion of this receptor (see below).

The suggestion that noradrenaline has higher potency at α_1D-adrenoceptors is confirmed in many pharmacological studies. Indeed, noradrenaline has been reported to have high potency (aorta: pEC50 of 8.15) [24] in tissues with a high level of α_1D-adrenoceptor messenger RNA (mRNA; aorta: 70–80 %; [89a]) and low potency (pEC50 of 6.32) [126] in the small mesenteric artery which had a high level of α_1A-adrenoceptor mRNA (75 %; [89a]).

In mouse mesenteric arteries, where, as discussed above, the predominant receptor is α_1A-adrenoceptor (see Fig. 4.2), prazosin potency (pK _B, −log M) was 8.8 in WT mice, but 9.6 in α_1B-/α_1D-adrenoceptor KO mice [91]. However, prazosin pK _B was 10.3 in α_1A-/α_1B-adrenoceptor KO mice [92]. In another study, prazosin pA ₂ was 9.92 in WT and 9.83 in α₁B KO, but 9.30 in α_1D KO and 9.43 in α_1B/α_1D KO [67]. These studies seem to agree that prazosin may be more potent in the mouse than in the rat, but there is no conclusive evidence that its potency is decreased by α_1DKO.

Piascik et al. (1995) [110] reported that the α_1A-adrenoceptor subtype plays a role in the tonic maintenance of blood pressure in the conscious rat, whereas the α_1B-adrenoceptor subtype participates in the response to exogenous agonists. In receptor KO mice, α_1A-adrenoceptor KO significantly reduced resting blood pressure in only two from four studies, and indeed a combined α_1A-/α_1B-adrenoceptor KO failed to affect basal blood pressure in two studies (see Table 4.1), α_1B-adrenoceptor KO did not affect resting blood pressure, but α_1D-adrenoceptor KOs significantly reduced resting blood pressure in three from three studies (Table 4.1). In KO mice lacking the α_1B-adrenoceptor subtype, there was no effect on basal blood pressure, but pressor responses to phenylephrine were significantly blunted [21]. However, overexpression of α_1B-adrenoceptors results in hypotension and cardiac hypertrophy, suggesting that effects seen in α_1B-adrenoceptor KO mice may not relate to direct blood pressure actions of α_1B-adrenoceptors (otherwise, overexpression would result in increased blood pressure) [151]. In the pithed rat, a component of the vasopressor nerve response, previously identified as α₂-adrenoceptor mediated based on the effects of the α₂-adrenoceptor antagonist yohimbine, has been reclassified as α_1D-adrenoceptor mediated [35]. Yohimbine was found to have unexpected α_1D-adrenoceptor potency. Hence, in the pithed rat, pressor nerve responses may involve both α_1A– and α_1D-adrenoceptors [35].

These results may suggest that α_1A– and α_1D-adrenoceptors and perhaps α_1B-adrenoceptors are involved in blood pressure control . However, the clearest result in terms of basal blood pressure is with the α_1D-adrenoceptor KO mouse: Blood pressure has been consistently reported to be significantly lowered in α_1D KO mice (see Table 4.1).

Following the identification of prejunctional α₂-adrenoceptors, evidence was presented, employing α₂-adrenoceptors antagonists and especially yohimbine, that α₂-adrenoceptors may be present on vascular smooth muscle and to mediate vasoconstrictor responses, in terms of actions both of exogenous agonists and of endogenous neurotransmitters [39, 40, 41]. More recently, the subtypes of α₂-adrenoceptor involved in these pressor responses to exogenous agonists were investigated and it was found that the predominant receptor was α_2A-adrenoceptor [54]. However, it has since been established that the component of the vasopressor nerve responses in the pithed rat found to be blocked by yohimbine are actually α_1D-adrenoceptor mediated [35]. Fortunately, there is indeed an α_2A-adrenoceptor-mediated component to pressor responses to exogenous agonists in the pithed rat [36].

The effects of α₂-adrenoceptor receptor KOs on resting blood pressure have been examined: The only change in resting blood pressure produced by α₂-adrenoceptor KO is an increase in blood pressure in α_2A-adrenoceptor KO mice (see Table 4.1). The major effect of KO of the α_2A-adrenoceptors is abolition of a central hypotensive action [102]. As has been elegantly shown using KO mice, α_2A-adrenoceptors are involved in the central hypotensive actions of α₂-adrenoceptor agonists and hence the biphasic transient pressor and prolonged depressor response to the α₂-adrenoceptor agonist UK14,304 in WT mice was altered to a transient (but more prolonged) pressor response in α_2A-adrenoceptor KO mice [87, 88]. Hence, the crucial α₂-adrenoceptor in terms of blood pressure control is the α_2A-adrenoceptor, involved presumably in tonic central control of blood pressure (although a component due to peripheral effect at prejunctional α_2A-adrenoceptors on sympathetic nerves innervating blood vessels cannot be ruled out).

There were no obvious haemodynamic effects produced by disruption of the α_2C-adrenoceptor [84]. However, the biggest mystery surrounds the α_2B-adrenoceptor. Knockout of the α_2B-adrenoceptor abolished the early transient pressor response to the α₂-adrenoceptor agonist dexmedetomidine and prolonged the depressor response suggesting that the early pressor response is α_2B-adrenoceptor mediated [84]. These effects were admittedly obtained with a single dose of α₂-adrenoceptor agonist. These blood pressure effects are systemic cardiovascular responses and do not allow for subtle effects on different vascular beds, nor do they prove categorically that the α_2A-adrenoceptor subtype has purely central actions or that the α_2B-adrenoceptor subtype is the only mediator of pressor responses. It is possible that α_2A-adrenoceptors mediate both central hypotension and peripheral hypertension, but the central actions dominate in conscious animals.

The receptor knockout studies would suggest that the α_2B-adrenoceptor and not the α_2A-adrenoceptor is involved in peripheral pressor responses or at least that the α_2B-adrenoceptor is the predominant receptor involved under the experimental conditions and with the chosen agonist. How do these findings on blood pressure in KO mice relate to the above studies in the pithed rat in which pressor responses were mediated by α_2A-adrenoceptors (see above)? The pithed rat results do not rule out the involvement of α_2B-adrenoceptors in pressor responses in pithed rats, but suggest that α_2A-adrenoceptors are dominant. The identification of α_2A-adrenoceptors relies on BRL 44408, but no reliable selective α_2B-adrenoceptor antagonist is available. The lack of really good selective α_2B– or indeed α_2C-adrenoceptor antagonists hinders pharmacological studies. We need more selective antagonists to answer these questions.

Hence, in general, it seems that the major contributors to exogenous-agonist-induced pressor responses in vitro and to exogenous-agonist-induced contractions in vitro are α_1A-, α_1D– and α_2A (or α_2B)-adrenoceptors. However, effects of exogenous agonists, while of importance in terms of pharmacological actions of drugs, may tell us little about what receptors are important in the physiological control of the vasculature. Hence, KO studies can be of use in clarification of this point.

In terms of postjunctional α₂-adrenoceptors in isolated tissues, there is relatively little evidence available as to subtypes mediating vascular contractions , given the relatively few isolated tissue preparations in which these receptors can be demonstrated. The human saphenous vein is a preparation in which it appears that the dominant adrenoceptors mediating contractions are α₂-adrenoceptors. In studies from this laboratory of side branches of the human saphenous vein, there appears to be a homogeneous population of α₂-adrenoceptors with no evidence for α₁-adrenoceptors : Yohimbine was more potent than prazosin; both yohimbine and prazosin produce parallel shifts in the noradrenaline concentration–response curve [see 122]; other α₁-adrenoceptor antagonists such as HV 723 [100] had relatively low potency; the α₁-adrenoceptor selective agonist phenylephrine had low potency; and the α₂-adrenoceptor selective agonist oxymetazoline had high potency [55]. Other studies of the human saphenous vein have reported that contractions are mediated predominantly by α₂-adrenoceptors [99, 128], or by α₁– and α₂-adrenoceptors [43, 114]. However, it is important to note that α₂-adrenoceptors identified employing yohimbine at a time before the functional identification of the α_1D-adrenoceptor must be treated with caution until selective α_1D-adrenoceptor antagonists are also examined: Yohimbine has actions at α_1D-adrenoceptors in concentrations only five- to tenfold less than those at which it has actions at α₂-adrenoceptors. Hence, some of these results have been reconsidered in terms of possible α_1D-adrenoceptor involvement: In the human saphenous vein, responses to exogenous noradrenaline have been shown to be α_2C– [55] and not α_1D-adrenoceptor mediated [24].

In the porcine palmer lateral vein and common digital artery, the low potency of prazosin suggests the presence of α_2A-adrenoceptors [12], and α_2A-adrenoceptors have also been reported in dog saphenous vein [86]. In the pig ciliary artery, the high potency of BRL44408 suggests the involvement of α_2A-adrenoceptors in contractions [143]. In a number of tissues, α₂-adrenoceptors contribute to a predominantly α₁-adrenoceptor-mediated response. In rat cremaster arterioles and venules, a component of the contractile response, based on the potency of BRL 44408, seems to be mediated by α_2A-adrenoceptors [83].

Initially, it was suggested that vascular α₂-adrenoceptors may be predominantly extrasynaptic and mediate responses to circulating catecholamines [79], but it was later found that contractions to nerve stimulation in the human saphenous vein are α₂-adrenoceptor mediated, based on high potency of yohimbine and low potency of prazosin [38]. The low potency of prazosin rules out an important role for α_1D-adrenoceptors in this response.