,nal of Cardio vascular Pharmacology
`~•H-558 © 1984 Raven Press, New York
`
`Alpha-Adrenoreceptor Subtypes in Blood Vessels:
`Physiology and Pharmacology
`
`S. Z. Langer and P. E. Hicks
`
`Department of Biology, Laboratoires d' Etudes et de Recherches Synthelabo, Paris, France
`
`Summary: A significant advance in the field of neuro(cid:173)
`transmission was made with the discovery of presynaptic
`release-modulating alpha-adrenoreceptors on noradren(cid:173)
`ergic nerve terminals. The concept of presynaptic mod(cid:173)
`ula1ion of noradrenaline release developed in parallel with
`Ihe pharmacological evidence for two subtypes of alpha(cid:173)
`adrenoreceptors as defined by a different profile of af(cid:173)
`finity and relative order of potencies for agonists and for
`anlagonists. The alpha 1-adrenoreceptor is stimulated
`preferentially by methoxamine and cirazoline and
`blocked selectively by prazosin or corynanthine. The
`~phai-adrenoreceptor is stimulated preferentially by ag(cid:173)
`onists such as clonidine, TL-99 , GHT-933 , and UK-
`14,304, and the responses mediated by these agonists are
`selectively blocked by the alphar adrenoreceptor antag(cid:173)
`onist idazoxan. In blood vessels , both the alpha 1- and the
`~phaz-adrenoreceptor subtypes are present postsynaptj-
`
`cally, where they mediate vasoconstriction, although the
`alpha 1-adrenoreccptor is the predominant receptor in vas(cid:173)
`cular smooth muscle. Presynaptically on noradrenergic
`nerve terminals , the stimulation of inhibitory alpha,-ad(cid:173)
`renoreceptors reduces the depolarization-evoked release
`of the transmitter. In most vascular beds , the alpha 1-ad(cid:173)
`renoreceptor is also the preferentially innervated sub(cid:173)
`type. In spontaneously hypertensive rats , smooth-muscle
`alpha2-adrenoreceptors mediate vasoconstrictor re(cid:173)
`sponses to exogenous noradrenaline and to sympathetic
`nerve stimulation to a greater extent than in the corre(cid:173)
`sponding normotensive Wystar-Kyoto rats , which may
`point to a pathophysiological role of these alpha2-adre(cid:173)
`noreceptors in hypertension . Key Words: Alpha 1-and
`alpha2 - adrenoreceptors-Noradrenaline- Noradrener(cid:173)
`gic neurotransmission-Presynaptic receptors -Dopa(cid:173)
`mine- Hypertension.
`
`Until approximately 10 years ago , it was gener(cid:173)
`ally accepted that alpha-adrenoreceptors repre(cid:173)
`sented a homogeneous population. The discovery
`of presynaptic alpha-adrenoreceptors and their role
`m the modulation of noradrenergic neurotransmis(cid:173)
`sion (1-3) provided the stimulus for the subclassi(cid:173)
`fication of alpha-adrenoreceptors into alpha1- and
`alphar subtypes. This subclassification developed
`as a result of the pharmacological differences be(cid:173)
`tween presynaptic (alpha2) and posts ynaptic
`(alpha1) adrenoreceptors (2).
`The original proposal for the subclassification of
`alpha-adrenoreceptors was based on pharmacolog(cid:173)
`ical differences observed using alpha-adrenore(cid:173)
`ceptor r.locking agents (4,5) on peripheral presyn(cid:173)
`aptic and posts ynaptic alpha-adrenoreceptors in
`
`the perfused cat spleen (2,4 ,5). Subsequent work in
`this field , using a number of selective agonist and
`antagonist drugs , has confirmed the existenc e of
`alpha 1- and alpha2-adrenoreceptor subtypes and al(cid:173)
`lowed a characterization of their location and func(cid:173)
`tion (3 ,6).
`In blood vessels, the presynaptic receptor, which
`inhibits the release of noradrenaline , corresponds
`to the alpha2-subtype , while postsynaptically, in
`smooth muscle, alpha1- as well as alphar adreno(cid:173)
`receptors are present a nd be-th are involved 'Nith
`the contractile process.
`The present article deals with the physiological
`and pharmacological relevance of the alpha -adre(cid:173)
`noreceptor subtypes in blood vessels under norma l
`conditions and in the hypertensive state.
`
`Address correspondenc e and reprint requests to Dr. La nger at De pa rtme nt of Biology L. E. R .S., 58, rue de la Glaciere, 7501 3 Paris,
`France.
`
`S547
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`

`S548
`
`S. Z. LANGER AND P. E. HICKS
`
`TABLE 1. Pharmacological basis for the
`subclassification of alpha-adrenoreceptors
`
`TABLE 2. Pharmacological basis for the
`subclassification of alpha-adrenoreceptors
`
`Order of selectivity for agonists
`
`Order of selectivity for antagonists
`
`Methoxamine
`Amidephrine
`Cirazoline
`Phenylephrine
`Noradrenaline
`Adrenaline
`Alpha-CHrnoradrenaline
`Dopamine
`6-F-noradrenaline
`Clonidine
`Para-aminoclonidine
`M-7
`UK-14304
`TL-99
`BHT-920
`BHT-933
`
`Alpha-adrenoreceptor agonists in their order of selectivity for
`the alpha 1- and alpha2-adrenoreceptor subtypes.
`M7 = [2,N,N-dimethylamino-5,6-dihydroxy-1,2,3,4-tetrahy(cid:173)
`dronaphthalene].
`TL99 = [2,N ,N-dimethylamino-6,7-dihydroxy-1,2,3,4-tetra(cid:173)
`hydronaphthalene].
`BHT920 = [2-amino-6-allyl-5 ,6, 7 ,8-tetrahydro-4H-thiazolo(cid:173)
`(4,5-d)azepine).
`BHT933 = [2-amino-6-allyl-4,5 , 7,8-tetrahydro-6H-oxazolo(cid:173)
`(5 ,4-d)azepine].
`UK 14304 = [5-bromo-6-(2-imidazolin-2-ylamino)-quinoxa(cid:173)
`line].
`
`AGONISTIC AND ANTAGONISTIC DRUGS
`ACTING ON ALPHA-ADRENORECEPTOR
`SUBTYPES: PHARMACOLOGICAL BASIS
`FOR RECEPTOR SUBCLASSIFICATION
`The relative order of selectivities of alpha-adre(cid:173)
`noreceptor agonists acting on alpha1- and alphar
`adrenoreceptor subtypes is shown in Table 1. Meth(cid:173)
`oxamine, amidephrine, and cirazoline, followed
`by phenylephrine, are among the most selective
`alpha1-adrenoreceptor agonists. Whereas noradren(cid:173)
`aline and adrenaline stimulate both subtypes of
`alpha-adrenoreceptors, the alpha-methyl and the 6-
`fluoro analogs of noradrenaline are preferential
`alpharadrenoreceptor agonists (Table 1).
`Among the preferential alpharadrenoreceptor ag(cid:173)
`onists which are not phenylethylamines, clonidine,
`para-aminoclonidine, BHT-920 , BHT-933, UK-
`14304, and the aminotetralines M-7 and TL-99 are
`the most widely used drugs (Table 1). It is of interest
`that in these series of compounds, M-7, TL-99, and
`BHT-920 are also agonists at the Di-receptor (7).
`Finally, although dopamine itself is a weak alpha(cid:173)
`adrenoreceptor agonist, it is also of interest that in
`several experimental models, dopamine preferen(cid:173)
`tially stimulates the alphar rather than the alpha1-
`adrenoreceptor subtype (8,9). Consequently, when
`the relative order of potency for dopamine, com(cid:173)
`pared with noradrenaline, on the alpha-adrenore(cid:173)
`ceptor is determined, it is important to differentiate
`
`J Cardiovasc Plwrmacol, Vol. 6 (Suppl. 4), /984
`
`Prazosin
`Corynanthine
`WB4I01
`Phentolamine
`Mianserin
`Tolazoline
`Piperoxan
`Yohimbine
`Rauwolscine
`Idazoxan
`
`Alpha-adrenoreceptor antagonists in their order of selectivit)
`for the alpha,- and alpharadrenoreceptor subtypes.
`WB4 IOI = (N-[2-(2,6-dimethoxyphenoxy)-ethyl] 1,4-benzo(cid:173)
`dioxane-2-methylamine).
`
`the corresponding values for the alpha,- and the
`alpharadrenoreceptor subtypes.
`In addition to the relative order of selectivity of
`the agonists, the alpha-adrenoreceptor antagonist
`have provided the most solid piece of evidence in
`support of the subclassification of the alpha-adre(cid:173)
`noreceptor. Prazosin is a highly 'selective, compel•
`itive alpha1-adrenoreceptor antagonist (Table 2),
`and corynanthine (stereoisomer of yohimbine), al(cid:173)
`though less potent than prazosin, is also rather se(cid:173)
`lective as a blocking agent at alpha1-adrenorecep(cid:173)
`tors. This is not the case for WB4101, which is a
`preferential alpha1-adrenoreceptor antagonist, but
`is less selective than the former two antagonists.
`Phentolamine blocks both the alpha 1- and the
`alpharadrenoreceptor subtypes, and there are three
`other antagonists which belong to the group of non(cid:173)
`selective alpha-adrenoreceptor blocking agent :
`mianserin, tolazoline, and piperoxan (Table 2) (al(cid:173)
`though in some species, e.g., dog, piperoxan may
`be more selective for the alpharadrenoreceptor).
`Both yohimbine and rauwolscine are preferential
`alpharadrenoreceptor antagonists, while idazoxan
`is at present the most selective alpha2-adrenore(cid:173)
`ceptor antagonist available, showing a 100-fold se(cid:173)
`lectivity ratio between the alphar and alphar
`adrenoreceptors under in vitro conditions. Some
`analogs of idazoxan have recently been reported to
`possess an even greater selectivity ratio for the
`blockade of the alphar over the alpha1-adrenore(cid:173)
`ceptor subtype (10).
`
`LOCATION AND FUNCTION OF ALPHA(cid:173)
`ADRENORECEPTOR SUBTYPES IN BLOOD
`VESSELS: DIFFERENCES BETWEEN
`VASCULAR BEDS
`The neuroeffector junction at the adventitial me(cid:173)
`dial border in vascular smooth muscle is shown
`schematically in Fig. l. The alpha-adrenoreceptor
`subtype located presynaptically on noradrenergic
`nerve terminals and mediating an inhibition of nor-
`
`Slayback Exhibit 1090, Page 2 of 12
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`

`

`ALPHA-ADRENORECEPTOR SUBTYPES
`
`S549
`
`-t
`1.
`-t
`
`.?.~
`...
`:
`\
`...
`'
`f1
`'
`',
`~
`7
`·•.# .A
`
`"2
`
`♦ • ••
`• •
`
`♦
`♦ 3.
`■
`
`-
`
`constriction -
`
`BLOOD VESSEL
`
`FIG. 1. Alpha-adrenoreceptor subtypes in blood vessels.
`Schematic representation of a noradrenergic varicosity of
`lhe post-ganglionic sympathetic neuron innervating vascular
`smooth muscle. Electrical stimulation (1) leads to the exo(cid:173)
`cytotic release (2) of noradrenaline (squares) that crosses the
`junctional cleft to occupy (3) vascular alpha1-adrenorecep(cid:173)
`tors (squares indent) mediating vasoconstriction. Neuronally
`released noradrenaline may act at prejunctional alpha2-ad(cid:173)
`renoreceptors to inhibit the quantity of transmitter subse(cid:173)
`quently released per impulse. Postjunctional alpha2-adre(cid:173)
`noreceptors mediate vasoconstriction and appear to be
`predominantly extrasynaptic. When postjunctional alpha2-
`adrenoreceptors are stimulated by exogenously adminis(cid:173)
`lered agonists or by circulating catecholamines, a vasocon(cid:173)
`
`strictor response is elicited . .
`
`adrenaline release corresponds to the alpharsub(cid:173)
`type of adrenoreceptor. It is generally accepted that
`the presynaptic alpharadrenoreceptor is the target
`for exogenously administered agonists which re(cid:173)
`duce transmitter release (e.g., clonidine), but the
`physiological role of the presynaptic alpharadre(cid:173)
`noreceptor in modulating noradrenaline release is
`still controversial (11) . Depending on the frequency
`of nerve impulses and on the duration of depolar(cid:173)
`ization of noradrenergic nerves, alpharadrenore(cid:173)
`ceptor antagonists enhance the overflow of nor(cid:173)
`adrenaline during nerve stimulation. This indicates
`that there is an operational negative feedback mech(cid:173)
`anism through which the released transmitter nor(cid:173)
`adrenaline can autoregulate its own release, once
`a threshold concentration of noradrenaline is
`achieved in the synaptic cleft (12).
`Postsynaptically, in the vascular smooth muscle,
`the alpha 1-adrenoreceptor is the predominant re(cid:173)
`ceptor mediating vasoconstriction, but there are
`
`also postsynaptic alpharadrenoreceptors which
`mediate vasoconstriction in arterial as well as in
`venous vascular smooth muscle (Fig. 1, Tables 3,4).
`With the exception of the cerebral vascular bed,
`where the postsynaptic alphai-adrenoreceptor ap(cid:173)
`pears to be the predominant receptor in some of the
`species examined (Table 4), in most other peripheral
`vascular beds it is either the alpha 1-adrenoreceptor
`that predominates or both postsynaptic alpha1-
`and alpharadrenoreceptors that are present (Ta(cid:173)
`bles 3,4).
`It is of interest that, in the renal vascular bed ,
`there are almost exclusively alpha 1-adrenorecep(cid:173)
`tors , which subserve vasoconstriction, whereas in
`the femoral and mesenteric vascular beds, both
`alpha 1- and alpharadrenoreceptors appear to me(cid:173)
`diate vasoconstriction (Tables 3,4). Not only do the
`relative proportions of postsynaptic alpha1- and
`alpha2-adrenoreceptors vary a s a function of the
`vascular bed, but within a given vascular bed it is
`also possible that the proportions of alpha1- and
`alphaz-adrenoreceptors vary with the diameter of
`the blood vessel. Additional studies are required to
`establish the presence as well as the density of post(cid:173)
`synaptic alpha 1- and alphaz-adrenoreceptors in the
`smooth muscle of arteries and veins as a property
`of the vascular bed and also in relation to the caliber
`of the blood vessels. Finally, as shown in Tables 3
`and 4, it appears that there may be considerable
`species variations in the relative contributions of
`alpha 1- and alpha2-adrenoreceptor subtypes to va(cid:173)
`soconstriction in the different vascular beds. There(cid:173)
`fore , one should be cautious in extrapolating results
`from one species to another.
`
`PREFERENTIAL NORADRENERGIC
`INNERVATION OF THE ALPHA(cid:173)
`ADRENORECEPTOR SUBTYPE IN
`BLOOD VESSELS
`A number of studies have been carried out under
`in vivo as well as in vitro experimental conditions ,
`in which the effectiveness of alpha-adrenoreceptor
`antagonists was compared with respect to blocking
`the responses to exogenous noradrenaline and to
`the endogenously released transmitter (13-23) .
`In all these studies, the alpha 1-selective adreno(cid:173)
`receptor antagonist prazosin was more effective in
`blocking the responses to noradrenaline released by
`nerve stimulation than in antagonizing the same
`end-organ responses induced by exogenous nor(cid:173)
`adrenaline (13-23). Similar results were obtained
`with the use of other alpha 1-selective antagonists
`(18). These results were interpreted as follows: ex(cid:173)
`ogenous noradrenaline can activate both alpha 1-
`and alpha2-adrenoreceptors mediating vasocon(cid:173)
`striction (Table 1, Fig. 1), whereas the transmitter
`released by nerve stimulation elicits vasoconstric-
`
`J Cardiovasc Pharm acol, Vol. 6 (Suppl. 4), / 984
`
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`

`S550
`
`S. Z. LANGER AND P. E. HICKS
`
`TABLE 3. Distribution of alpha-adrenoreceptor subtypes in various vascular beds in vivo
`
`Vascular bed
`
`Species
`
`Experiments
`
`Alpha-receptor
`subtype
`
`References
`
`Whole animal
`Blood pressure
`
`Coronary
`circulation
`Thoracic/abdominal
`circulation
`
`Renal
`vasculature
`
`Femoral/hindlimb
`circulation
`
`Dog
`
`Cat
`Rabbit
`
`Rat
`Dog
`
`Dog
`
`Cat
`
`Dog
`Cat
`Rat
`
`Dog
`Dog
`Cat
`Rabbit
`Rat
`
`alpha / al pha2
`alpha/alpha2
`alpha/alpha2
`alpha/alpha2
`alpha/alpha2
`alpha1/alpha2
`
`alpha/alpha2
`
`alpha 1/alpha2
`
`alpha 1/alpha2
`alpha 1
`alpha 1
`
`alpha 1
`alpha/ alpha 1
`alpha/alpha2
`alpha/alpha2
`alpha/ alpha2
`
`(14, 15)
`(34)
`(35)
`(36)
`(37)
`(38-40)
`
`(4 I)
`
`(9)
`
`(21)
`(8 ,42)
`(21)
`
`(43)
`(13, 14,20,42)
`(44)
`(20)
`(45)
`
`Anesthetized
`Pithed
`Anesthetized
`Pithed
`Conscious
`Pithed
`Circumflex
`Coronary blood flow
`Mesenteric blood
`flow
`Mesenteric blood
`flow
`Renal blood flow
`Renal blood flow
`Renal blood flow
`(microspheres)
`Femoral blood flow
`Forelimb blood flow
`Femoral blood flow
`Hindlimb blood flow
`Hindquarter
`perfusion
`Hindlimb
`Forearm blood flow
`
`alpha 1/alpha2
`(46)
`alpha/ alpha2
`(46)
`alpha/ alpha2
`(47)
`The references quoted refer to the positive identification of postsynaptic alpha 1- or alpha2-adrenore-
`ceptor subtypes in these vascular beds in vivo.
`
`Man
`
`tion through the activation of alpha 1-adrenorecep(cid:173)
`tors (14,15). This point is exemplified in vitro in
`Sprague-Dawley rat-tail arteries (Fig. 2), since both
`prazosin and the alpha1-selective diastereoisomer of
`yohimbine, corynanthine (19,24), are particularly
`potent and effective antagonists of the response
`elicited by electrical field stimulation (for prazosin
`a concentration of only I nM virtually abolished the
`vasoconstrictor response to sympathetic nerve
`stimulation (Fig. 2). In the vascular beds examined
`thus far, it appears that the alpha1-adrenoreceptor
`is the preferentially innervated receptor, and it has
`been demonstrated that the neuronally mediated va(cid:173)
`soconstriction is exquisitely sensitive to blockade
`by prazosin (14,15,18,20,21).
`It is therefore possible that the alpha1-adrenore(cid:173)
`ceptor predominates in the adventitial-medial
`border, where most of the noradrenergic nerve ter(cid:173)
`minals are present, whereas the postsynaptic
`alpharadrenoreceptors are located mainly near the
`intima (Fig. 1) and therefore may be the target of
`circulating catecholamines (16). However, one
`should be cautious in generalizing about this ar(cid:173)
`rangement in blood vessels, because the preferen(cid:173)
`tial noradrenergic innervation of the alpha 1-adre(cid:173)
`noreceptor may also vary among vascular beds and
`with the diameter of the blood vessels. In addition,
`there may be differences between arteries and veins
`with respect to the alpha-adrenoreceptor subtype
`which is preferentially innervated; studies con(cid:173)
`cerning this question are presently underway in sev(cid:173)
`eral laboratories.
`In recent years, a number of electrophysiological
`
`J Cardiovasc Pharmacol, Vol. 6 (Suppl. 4), 1984
`
`studies have been carried out on arteriolar prepa(cid:173)
`rations, in which the excitatory junctional poten(cid:173)
`tials (EJPs) have been recorded in response to low(cid:173)
`frequency stimulation of the sympathetic nerves to
`the blood vessel (25,26). Although the EJPs are
`thought to be mediated by release of noradrenaline
`in these vessels, the electrophysiological responses
`remain resistant to blockade with all alpha-adre(cid:173)
`noreceptor antagonists (25-27). It is clearly pre(cid:173)
`mature to ascribe these phenomena to a new type
`of receptor [e.g., gamma-receptor (25)] in the ab(cid:173)
`sence of an antagonist which blocks the EJPs (27).
`Nevertheless, the phenomenon should not be dis(cid:173)
`missed, since these determinations represent a
`physiological measurement of the transmitter-re(cid:173)
`ceptor interaction, although it does not seem to in(cid:173)
`volve alpha-adrenorecepLors.
`
`CONTRIBUTION OF POSTSYNAPTIC
`ALPHAi- AND ALPHA2-ADRENORECEPT0RS
`TO V ASOCONSTRICTOR RESPONSES IN
`SPONTANEOUSLY HYPERTENSIVE RATS
`Postsynaptic alpha,- and alpharadrenoreceptors
`mediate vasoconstriction in both hypertensive rats
`[spontaneously hypertensive (SHR) and DOCA(cid:173)
`salt] and in normotensive controls (WKY) (28).
`Under in vivo conditions, in pithed SHR or WKY,
`the intravenous administration of alpha 1-selective
`agonists, such as phenylephrine, or the alpha2-se(cid:173)
`lective agonists (TL-99) (29) (Fig. 3), and BHT-993
`(30) causes similar increases in blood pressure, with
`
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`ALPHA-ADRENORECEPTOR SUBTYPES
`
`S551
`
`TABLE 4. Distribution of alpha-adrenoreceptor subtypes in isolated blood vessels
`
`Vascular bed
`
`Species
`
`Blood vessels
`
`Cerebral circulation
`
`Dog
`
`Extracranial
`circulation
`
`Coronary
`circulation
`J'horacic/abdominal
`circulation
`
`Renal vasculature
`
`Femoral/hindlimb
`circulation
`
`Cat
`Bovine
`Monkey
`Man
`Cat
`Dog
`Rabbit
`Dog
`Bovine
`Dog
`
`Man
`
`Rabbit
`
`Cat
`
`Rat
`
`Dog
`Rat
`
`Dog
`
`Rabbit
`
`Rat
`
`Man
`
`Basilar artery
`Middle cerebral
`Middle cerebral
`Pia! arteries
`Middle cerebral
`Middle cerebral
`Lingual artery
`Jugular vein
`Perfused ear artery
`Coronary artery
`Coronary artery
`Mesenteric artery
`Mesenteric vein
`Splenic artery
`Vena cava
`Portal vein
`Femoral artery
`Femoral vein
`Mesenteric/jejunal arteries
`Aorta
`Pulmonary artery
`Portal vein
`(longitudinal muscle)
`Mesenteric artery
`Perfused spleen
`Mesenteric bed
`Thoracic aorta
`
`Portal vein
`(longitudinal muscle)
`Portal vein
`(phasic activity)
`Renal vein
`Perfused kidney
`Renal artery
`Femoral artery
`Femoral vein
`Saphenous vein
`Saphenous vein
`Saphenous artery
`Tail artery (SHRSD)
`Tail artery (WKY)
`Digital arteries
`Metacarpal veins
`Umbilical arteries
`
`Alpha-receptor
`subtype
`
`Alphaifalpha1
`Alpha2
`Alpha2
`Alpha2 >> alpha 1"
`Alpha 1
`A1pha 1
`Alpha 1 > alpha2
`Alpha 1 > alpha2
`Alpha 1/alpha2
`Alpha 1
`Alpha1 >> alpha{
`Alpha / alpha2
`Alpha1/alpha2
`Alpha 1
`alpha1
`Alpha 1
`Alpha 1
`Alphaifalpha1
`Alpha 1
`Alpha 1
`Alpha 1
`
`Alpha 1
`Alpha 1
`Alpha / alpha2
`Alpha 1 >> alpha2
`Alpha 1
`Alpha 1 >> alpha2
`
`Alpha 1
`
`Alpha / alpha2
`Alpha 1
`Alpha 1
`Alpha 1
`Alpha 1
`Alpha / alpha2
`Alpha/alpha2
`Alpha 1/alpha2
`Alpha 1
`Alpha 1/alpha2
`Alpha 1
`Alpha/
`Alpha/ alpha/
`No evidence found
`
`References
`
`(48)
`(49)
`(50,51)
`(52)
`(49)
`(49)
`(51 )
`(53)
`(54)
`(55-57)
`(58)
`(57)
`(53)
`(59)
`(53)
`(53)
`(61 )
`(61 )
`(60)
`(62)
`(63)
`
`(62)
`(5 I)
`(64)
`(65)
`(66)
`(67)
`
`(66,68)
`
`(68)
`(53)
`
`(59)
`(59)
`(69,70)
`(71 )
`(72)
`(22,23 ,75)
`(22,23)
`(60,73)
`(73)
`(74)
`
`• Experiments on receptor binding.
`b Alpha-receptor subtype may not conform with classical alpha 1- or alpha2-adrenoreceptor subtype.
`c Hicks , unpublished observations.
`The references quoted refer to the positive identification of postsynaptic alpha 1- or alphaz-adrenoreceptor
`subtypes in these isolated blood vessels.
`
`no apparent difference in sensitivity between the
`normotensive and the hypertensive rats (Fig. 3). In
`this study, both phenylephrine and noradrenaline
`caused greater maximal vasoconstrictor responses
`in SHR, which is probably related to the adaptive
`tructural changes (increased wall/lumen ratio)
`known to occur in hypertension (31), since similar
`effects were observed with angiotensin II (28). Fur(cid:173)
`thermore, in pithed rats, there was no apparent
`change in the potency of yohimbine to block alphar
`adrenoreceptor-mediated vasoconstriction (28) in
`SHR as compared with WKY.
`Despite the overwhelming evidence for the pres-
`
`ence in vivo of post-synaptic alphar adrenorecep(cid:173)
`tors which mediate vasoconstriction , it has been
`particularly difficult to demonstrate their presence
`in arterial smooth muscle in vitro. It is most likely
`that only arteriolar constriction, and not an influ(cid:173)
`ence on the venous circulation, accounts for the
`changes in blood pressure, blood flow, and vascular
`resistance (32) which are observed in vivo in re(cid:173)
`sponse to alpharadrenoreceptor agonists.
`In vitro, the perfused/superfused tail artery of
`SHR appears to contain, in addition to the well(cid:173)
`established alpha 1-adrenoreceptor, a population of
`postsynaptic alpha2-adrenoreceptors which me-
`
`J Cardiovasc P/Jarmacol, Vol. 6 (Suppl. 4/, /984
`
`.. .. .,
`,,.
`
`!.,
`t'
`
`:, . 4
`
`Slayback Exhibit 1090, Page 5 of 12
`Slayback v. Eye Therapies - IPR2022-00142
`
`

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`S552
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`S. Z. LANGER AND P. E. HICKS
`
`PRAZOSIN
`
`IDAZOXAN
`
`CORYNANTHINE
`
`3
`
`10
`
`30
`
`10
`3
`30
`Frequency of stimulotion (Hz}
`
`3
`
`10
`
`30
`
`FIG. 2. Antagonism produced by
`alpha-adrenoreceptor antagonists ol
`the vasoconstrictor responses in•
`duced by electrical field stimulation in
`the isolated perfused rat-tail artery
`(Sprague-Dawley). Ordinate, peak
`contractile responses to electrical
`stimulation expressed as percentage
`of the maximal response to noradren(cid:173)
`aline (NA). Abscissae , frequency ol
`stimulation (Hz), monophasic square(cid:173)
`wave pulses of 0.3 ms duration. Su•
`pramaximal voltage. Left panel, (I )
`controls ; prazosin (.6.)1 nM, (♦)100
`nM. Middle panel, (e) controls; ida(cid:173)
`zoxan (♦)100 nM, (■)1 µM, (.6.)10 µ.M.
`Right panel, (e) controls; corynan(cid:173)
`thine (■) 1 µM, (♦)10 µM. Incubation
`time with each antagonist was 20 min.
`Cocaine (4 µM) and propranolol (1
`µM) were present in the Krebs' perfu(cid:173)
`sion medium. Vertical bars indicate
`mean ± SEM for at least four experi(cid:173)
`ments. Data taken from reference 75.
`
`diate vasoconstriction in this vessel (22,23). Re(cid:173)
`sponses induced by noradrenaline and the aJphar
`adrenoreceptor agonist TL-99 are antagonized by
`the selective alpharadrenoreceptor antagonist ida(cid:173)
`zoxan (22,23) at concentrations which do not an(cid:173)
`tagonize responses induced by the alpha1-adreno(cid:173)
`receptor agonist methoxamine (Fig. 4). This antag(cid:173)
`onistic effect of low concentrations of idazoxan
`has not been observed in tail arteries from WKY
`(Fig. 4.)
`Similarly to the vasoconstrictor responses pro(cid:173)
`voked by exogenous noradrenaline, those mediated
`
`by electrical field stimulation in SHR tail arteries
`were significantly greater than those obtained in
`WKY arteries (22,23) (Fig. 5). The alpha1-adreno(cid:173)
`receptor is the predominant subtype in both SHR
`and WKY tail arteries. Although prazosin was a
`very potent antagonist of the responses to sympa(cid:173)
`thetic nerve stimulation, a significant antagonism of
`the neuronally-mediated vasoconstriction could
`also be demonstrated in tail arteries of SHR with
`idazoxan (Fig. 5) at concentrations which do not
`block alpha 1-adrenoreceptor-mediated constriction.
`This profile of antagonism, with low concentrations
`
`FIG. 3. Vasoconstrictor responses in(cid:173)
`duced by alpha-adrenoreceptor ago(cid:173)
`nists in pithed (SHA) or normotensive
`WKY. SHA and WKY were selected on
`the basis of systolic blood-pressure
`measurements (tail-cuff method in
`conscious rats). SHA were used if the
`systolic pressure exceeded 185 mm
`Hg; WKY were only used if systolic
`blood pressure was less than 140 mm
`Hg. Basal diastolic blood pressures of
`these rats 30 min after pithing were
`not significantly different (SHA = 39
`± 4 mm Hg, n = 27; WKY = 42 ± 3
`mm Hg, n = 28). Ordinate, increase in
`diastolic blood pressure (mm Hg). Ab(cid:173)
`scissae, log-dose agonists (µg/kg i.v.).
`Left panel, phenylephrine. Middle
`panel, noradrenaline (NA). Right
`panel, TL-99. •-•. response curves
`obtained in WKY; 0---0, response
`curves obtained in SHA. * p < 0.05,
`significantly different from SHA data.
`Vertical bars indicate mean ± SEM.
`Data taken from reference 28.
`
`J Cardiovasc Pharmacol, Vol. 6 (Suppl. 4), /984
`
`Q-Q
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`I
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`
`0
`
`10
`100
`Phenylcphrine (µg/ kg i.v.)
`
`0.01
`
`0.1
`NA (µg/ kg i.v.)
`
`10
`
`0.1
`10
`TL -99 (µg/ kg i.v.)
`
`Slayback Exhibit 1090, Page 6 of 12
`Slayback v. Eye Therapies - IPR2022-00142
`
`

`

`ALPHA-ADRENORECEPTOR SUBTYPES
`
`S553
`
`SHR
`
`I
`I
`
`NA
`
`I
`I
`
`I
`
`Fr
`-d" METHOXAMINE
`
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`u
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`_g
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`0
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`WKY
`
`METHOXAMINE
`
`FIG. 4. Antagonistic effects of ida-
`zoxan against vasoconstrictor re-
`sponses induced by alpha-adrenore-
`ceptor agonists in perfused tail ar-
`teries from SHR or WKY normotensive
`controls. Schild plots for idazoxan
`against TL-99 (L - L), noradrenaline
`(NA) (e-e), or methoxamine (0-□)
`in SHR tail arteries (left panel) or WKY
`tail arteries (right panel). Ordinate, log
`(concentration ratio - 1 ). Abscissae, log
`molar concentration of idazoxan. Ver-
`tical bars indicate mean ± SEM. n =
`at least four experiments for each
`point. Note that idazoxan was signifi-
`cantly more potent as an antagonist of
`responses evoked by TL-99 or nor-
`adrenaline than against methoxamine
`in SHR tail arteries, but was equipo-
`tent against these three agonists in
`WKY tail arteries.
`
`-8
`
`.7
`-5
`-6
`Log M [ldazoxan]
`
`-4
`
`-8
`
`,-5
`-7
`-6
`Log M [ldazoxan]
`
`-4
`
`of idazoxan, was not demonstrated for the vaso(cid:173)
`constriction elicited by nerve stimulation in WKY
`tail arteries (Fig. 5).
`ldazoxan has been shown to act as a partial ag(cid:173)
`onist at alpharadrenoreceptors in perfused rabbit(cid:173)
`ear arteries (33); however, we have now demon(cid:173)
`strated that low concentrations of idazoxan, which
`inhibit end-organ responses to field stimulation, do
`not reduce the electrically evoked overflow of 3H(cid:173)
`noradrenaline from SHR tail arteries (Langer and
`Hicks, unpublished observations). We therefore
`conclude that, in the SHR-tail artery preparation, a
`significant population of postsynaptic alpharad(cid:173)
`renoreceptors can be demonstrated. Furthermore,
`in hypertensive animals, these alpharadrenorecep(cid:173)
`tors may also be activated by endogenously re(cid:173)
`leased noradrenaline to elicit vasoconstriction. The
`difficulties in demonstrating changes in post-syn(cid:173)
`aptic alphaz-adrenoreceptor-mediated vasoconstric-
`
`tion in vivo (involving the total circulation) empha(cid:173)
`size the necessity to evaluate these mechanisms in
`localized vascular beds, where the postsynaptic
`alpha 1- and alpharadrenoreceptor subtypes are
`likely to play an important role in the modulation
`of regional blood flow. Such studies in regional vas(cid:173)
`cular beds in hypertensive models may provide
`valuable information on the role of vascular alphar
`adrenoreceptors in the development or mainte(cid:173)
`nance of hypertension.
`
`CONCLUSIONS
`There is now a wealth of experimental evidence
`in support of the subclassification of alpha-adren(cid:173)
`oreceptors into alpha 1- and alpharsubtypes.
`The alpha 1-adrenoreceptor subtype, linked to va(cid:173)
`soconstriction, is the predominant postsynaptic re(cid:173)
`ceptor in vascular smooth muscle and in most blood
`
`150
`
`a,
`I
`E
`i
`~ ,
`: 100
`~
`a.
`C
`0 -~ ,
`
`'t
`11>
`a. 50
`C
`
`11>
`~
`0
`~
`u
`C
`
`0
`
`,0.3
`
`0.3
`3
`Electrical stimulation (Hz)
`
`3
`
`FIG. 5. Blockade by the alpha-adrenoreceptor
`antagonists idazoxan or prazosin of the re(cid:173)
`sponses induced by electrical field stimulation
`of the isolated perfused tail artery preparation
`from SHR, or normotensive controls WKY. Or(cid:173)
`dinate, increase in perfusion pressure (mm Hg).
`Abscissae, frequency of stimulation (Hz), mono(cid:173)
`phasic square-wave pulses of 0.3 ms duration,
`supramaximal voltage. Control (C) frequency(cid:173)
`response curves (e-e). Antagonistic effects of
`idazoxan (IDZ) ( ■-■ ; 10 nM; 20 min preincu(cid:173)
`bation , n = 6), or prazosin (PRZ) (&-&; 10 nM;
`20 min preincubation, n = 6) in SHR (left panel)
`and WKY (right panel). Vertical bars indicate
`mean ± SEM. *p < 0.05; **p < 0.01 , significantly
`different from control data. Note that idazoxan
`(10 nM) significantly reduced the responses to
`field stimulation in SHR at 0.3 and 1 Hz, whereas
`it did not affect the corresponding responses in
`WKY normotensive tail arteries. Data taken from
`reference 23.
`
`J Cardiovasc Pharmacol. Vol. 6 (Suppl. 4), 1984
`
`Slayback Exhibit 1090, Page 7 of 12
`Slayback v. Eye Therapies - IPR2022-00142
`
`

`

`S554
`
`S. Z. LANGER AND P. E . HICKS
`
`vessels, appears to be the preferentially innervated
`receptor. This subtype seems to be located predom(cid:173)
`inantly in the medial adventitial border.
`The postsynaptic alpha2-adrenoreceptor sub(cid:173)
`type also mediates contraction of vascular smooth
`muscle and appears to be located close to the intima
`of blood vessels, where it may be the target of cir(cid:173)
`culating catecholamines rather than neuronally re(cid:173)
`leased noradrenaline.
`The relative proportions of postsynaptic alpha1-
`and alphaz-adrenoceptors in blood vessels vary
`with the vascular bed, the arterial or venous sec(cid:173)
`tions, and the diameter of the blood vessel. In ad(cid:173)
`dition, species differences apparently exist in this
`proportion in certain vascular beds.
`In SHR there is a postsynaptic alpha2-adreno(cid:173)
`receptor-mediated component of vasoconstriction
`to both exogenous and endogenous noradrenaline.
`On the other hand, alphaz-adrenoreceptors in vas(cid:173)
`cular smooth muscle may contribute significantly
`less to vasoconstriction in normotensive WKY rats.
`These results suggest that postsynaptic vascular
`alpha2-adrenoreceptors mediating vasoconstriction
`may play an important role in the pathophysiology
`of hypertension and contribute to the increased vas(cid:173)
`cular reactivity to noradrenaline observed in hy(cid:173)
`pertensive states.
`At a presynaptic level, inhibitory alphaz-adreno(cid:173)
`receptors are present on noradrenergic nerve ter(cid:173)
`minals, and their activation leads to a reduction in
`the output of noradrenaline during nerve stimula(cid:173)
`tion. The physiological role of presynaptic inhibi(cid:173)
`tory alphaz-adrenoreceptors in the modulation of
`noradrenergic neurotransmission depends on the
`frequency and duration of depolarization and on the
`presence of a certain threshold concentration of
`noradrenaline in the synaptic cleft. From the phar(cid:173)
`macologic