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Autoradiographic localization of α-adrenoceptors, muscarinic acetylcholine receptors and opiate receptors in the heart
European Journal of Pharmacology, 127 (1986) 279-282
279
Elsevier Short communication
AUTORADIOGRAPHIC LOCALIZATION OF a-ADRENOCEPTORS, MUSCARINIC A C E T Y L C H O L I N E R E C E P T O R S AND O P I A T E R E C E P T O R S IN T H E H E A R T MICHAEL R. DASHWOOD * and K. MICHAEL SPYER Department of Physiology, Royal Free Hospital School of Medicine, Rowland Hill Street, London NW3 2PF, England
Received 23 June 1986, accepted 24 June 1986
M.R. DASHWOOD and K.M. SPYER, Autoradiographic localization of a-adrenoceptors, receptors and opiate receptors in the heart, European J. Pharmacol. 127 (1986) 279-282.
muscarinic acetylchofine
Using in vitro autoradiography the distribution of [3H]rauwolscine, [3H]prazosin and [3H]quinuclidinyl benzilate binding sites has been demonstrated in cardiac tissue taken from the cat and rat. A similar distribution of both a 1- and a2-adrenoceptor sites was seen but the distribution of muscarinic acetylcholine sites was markedly different. a-Adrenoceptors were present predominantly in ventricular muscle whereas muscarinic acetylcholine receptors exhibited a greater density in atrial tissue compared to ventricular muscle. Opiate receptors were absent from cardiac tissue. Muscarinic acetylcholine receptors
a-Adrenoceptors
1. Introduction
a-Adrenoceptor, muscarinic acetylcholine receptor and opiate receptor agonists all affect heart rate when administered intravenously. The site of action of these compounds may be directly on the myocardium, mediated centrally or reflexly as a consequence of their effects on arterial blood pressure. Both muscarinic acetylcholine and al-adrenoceptors have been identified in membrane preparations of cardiac muscle (Guicheney and Meyer, 1981; N e d o m a et al., 1986) and eq-adrenoceptors have been shown to exist in the heart using autoradiography (Dashwood, 1983). In the present study we have used the in vitro autoradiographic technique of Young and Kuhar (1979) to localize a 1- and a2-adrenoceptors, muscarinic acetylcholine and opiate receptors in the heart of cat and rat.
* To whom all correspondence should be addressed. 0014-2999/86/$03.50 © 1986 Elsevier Science Publishers B.V.
Ventricular muscle
Autoradiography
Atria
2. Materials and m e t h o d s
Male and female Sprague-Dawley rats (RFHSM-derived, body weight around 250 g) and male and female cats (body weight around 3 kg) were used for this study. Animals were anaesthetized with sodium pentobarbitone (80 rng/kg i.p.) and perfused intracardially with ice-cold 100 m M phosphate buffer containing 0.32 M sucrose (pH 7.4) and lightly fixed with 0.1% formaldehyde solution in phosphate-sucrose buffer. After fixation the heart was removed, rinsed in buffer and mounted in Ames O.C.T. compound onto a cryostat chuck. Serial 20/~m longitudinal sections were taken through the heart at - 2 0 ° C which were thaw-mounted onto gelatinized microscope slides, allowed to dry and stored either overnight at - 2 0 ° C or for up to 4 weeks at - 7 0 ° C . Paired slides were then taken and incubated for 60 min at 4 ° C in 4 nM [3H]naloxone (specific activity 55 C i / m m o l ) made up in 170 m M Tris HC1 buffer containing 100 m M NaC1 (pH 7.4), 60 min at 4 ° C in 5 nM [3H]prazosin (specific activity 33 C i / mmol) made up in 170 m M Tris HC1 buffer (pH
280 7.4) or for 120 min at 4 ° C in 2 nM [3H]rauwolscine (specific activity 80 C i / m m o l ) and 1 nM [3H]quinuclidinyl benzilate (QNB) (specific activity 38 C i / m m o l ) both made up in 100 m M phosphate buffer (pH 7.4) in order to identify opiate, al-adrenoceptors, a2-adrenoceptors and muscarinic acetylcholine receptors respectively. After incubation sections were washed twice in the appropriate buffers before being blown dry in a cold stream of air, placed into 24 x 30 cm X-ray cassettes and apposed to LKB Ultrofilm for 4-6 weeks at 4°C. The degree of non-specific binding to tissue sections was established in the presence of 1/~M naloxone or morphine (opiates), 1-10/~M phentolamine (a~- and a2-adrenoceptors ) and 1 /~M carbachol (muscarinic acetylcholine receptors). After exposure the LKB Ultrofilm was developed in Kodak D19 developer (undiluted) for 5 min at 20°C, fixed in Ilfospeed fixer (diluted 1 to 4 with distilled water) for 5 min at 20°C and rinsed for at least 20 min at 20 °C before drying. Autoradiographs were visually inspected and photographed, where appropriate, using a Nikon Macro system. All tritiated ligands were purchased from Amersham International.
[3H]QNB binding to cardiac muscle in general which represents the presence of muscarinic acetylcholine receptor sites. Seventy three percent of this binding was displaced by incubating sections in the presence of 1 /~M carbachol. There was, however, a regional variation in this binding with the [3H]QNB binding being much greater in ventricular muscle than both a 1- and a2-adrenoceptors. There was a most striking muscarinic acetylcholine receptor presence in the atria (figs. 1A and 2A).
3. Results There were no qualitative differences in the distribution of the various receptors in the heart of both cats and rats. Opiate receptors, indicated by [3H]naloxone binding, were absent from all regions of the heart (figs. 1B and 2B). Both cqand a2-adrenoceptors ([3H]prazosin and [3H]rauwolscine binding) exhibited a fairly high and variable distribution over ventricular muscle with a lower degree of binding to atrial tissue (figs. 1C-D and 2C-D). This binding was to specific a 1- and a2-adrenoceptor sites since both [3H]prazosin and [3H]rauwolscine were displaced to a marked degree (about 80%) in those sections incubated in the presence of 1/~M unlabelled phentolamine. It is interesting to note that there was a high degree of [ 3H]prazosin binding to the inner surface of the aorta (arrowed in fig. 2C) whereas there was no [3H]rauwolscine binding in the adjacent section (fig. 2D). There was an extremely high degree of
Fig. 1. Autoradiographic localization of receptors in consecutive sections of rat heart. (A) Section incubated in 1 nM [3H]QNB. There is a very dense binding at the atria (A) with a lower distribution over the ventricular muscle (V). (B) Section incubated in 4 nM [3H]naloxone. There is no [3H]naloxone binding to cardiac tissue. (C) Section incubated in 5 nM [3H]prazosin. There is a generalized moderate distribution of binding over the ventricular muscle with a reduced binding at atrial regions. (D) Section incubated in 2 nM [3H]rauwolscine. There is a generalized moderate distribution of binding over the ventricular muscle with a slightly reduced binding at atrial regions. Scale bar = 2.5 mm.
281
Fig. 2. Autoradiographic localization of receptors in consecutive sections of cat heart. (A) Section incubated in 1 nM [3H]QNB. There is a very dense binding to ventricular muscle (V) accompanied by extremely dense binding at atrial areas (A). (B) Section incubated in 4 nM [3H]naloxone. There is no binding to cardiac tissue. (C) Section incubated in 5 nM [3H]prazosin. There is a generalized high binding to ventricular muscle with a much weaker binding to the atria. There is a strong binding of [3H]prazosin to the internal surface of the aorta (arrowed). (D) Section incubated in 2 mM [3H]rauwolscine. There is a generalized high binding to ventricular muscle with a weaker binding to the atria. Scale bar = 5 mm.
4. Discussion O p i a t e receptor, muscarinic acetylcholine receptor and ct-adrenoceptor agonists all elicit effects on the heart when administered intravenously. These effects m a y be mediated centrally since receptors for these c o m p o u n d s have been identified in brain areas associated with cardiovascular control (Young and Kuhar, 1979; W a m s ley et al., 1981; Unnerstall et al., 1984).
The purpose of the present autoradiographic study was to establish whether opiate receptors, muscarinic acetylcholine receptors and a-adrenoceptors exist in the heart of the cat and rat where direct a c t i o n s m a y be elicited. There was no [3H]naloxone binding to cardiac tissue of either species (figs. 1B and 2B). Heart sections were incubated in parallel with brain and spinal cord tissue which exhibited areas of high [3H]naloxone binding in, for example, the nucleus of the tractus solitarii and substantia gelatinosa of the thoracic spinal cord ( D a s h w o o d et ak, 1985). This result would indicate that opiate receptors are absent from cardiac tissue and that opiate-induced bradycardia may be due, in part, to an effect on receptors within the dorsomedial brainstem (see D a s h w o o d et al., 1985). There was a fairly dense distribution of both a 1 ([3H]prazosin binding)and a 2 ([3H]rauwolscine binding)-adrenoceptors on the heart. The distribution was similar in tissue from both cat and rat (figs. 1 and 2). The highest density of a-adrenoceptors was in ventricular muscle with a reduced binding at atrial regions (figs. 1C-D and 2C-D). Subtle variations in receptor density m a y well have been present across sections of ventricular tissue. There certainly appears to be a higher degree of [3H]QNB binding at the epicardium than endocardium. However there was no obvious increase in any receptor density at specialized areas of the heart such as the SA or A V node. Subsequent densitometric analysis of these autoradiographs m a y reveal subtle regional variations in receptor distribution. Figure 2 A - D illustrate four consecutive 20 /~m sections through the heart. It is interesting to note that in this particular plane a strong binding of [3H]prazosin is evident on the internal surface of the aorta whereas [3H]rauwolscine binding is absent (compare fig. 2C and D). This result is in agreement with the recent data of D a s h w o o d and Jacobs (1985) who showed a presence of al-adrenoceptors on the rat aorta but an absence of a 2adrenoceptors. Although there is considerable evidence for the presence of a~-adrenoceptors in cardiac muscle (Guicheney and Meyer, 1981), there is little substantial evidence for an a2-adrenoce ptor presence. According to the review of W a t a n a b e et al. (1982) the majority of ligand binding studies
282 carried out in c a r d i a c muscle have been with [3H]dihydroergocryptine, an a - a d r e n o c e p t o r antagonist, which will n o t d i s c r i m i n a t e between a 1a n d a2-subtypes. T h e most striking b i n d i n g o b s e r v e d in this s t u d y was that of [ 3 H ] Q N B to c a r d i a c tissue. T h e b i n d ing to ventricular muscle was stronger than that of [3H]prazosin a n d [3H]rauwolscine ( c o m p a r e figs. 1A a n d 2 A with figs. 1 C - D a n d 2 C - D ) but, whereas a - a d r e n o c e p t o r s were a l m o s t a b s e n t from atrial regions there was an e x t r e m e l y dense m u s c a r i n i c acetylcholine r e c e p t o r presence at this level (figs. 1A a n d 2A). These results not o n l y c o n f i r m results from r e c e p t o r b i n d i n g studies using h o m o g e n a t e prep a r a t i o n s of c a r d i a c tissue for a l - a d r e n o c e p t o r s a n d m u s c a r i n i c acetylcholine receptors (Guicheney a n d Meyer, 1981; N e d o m a et al., 1986) b u t also give an i n d i c a t i o n of the gross differential d i s t r i b u t i o n of such receptors across the heart. T h e r e are some i n d i c a t i o n s of the functional significance of these receptors. It is recognized that vagal efferent s t i m u l a t i o n causes a p r o f o u n d b r a d y c a r d i a which is b l o c k e d b y atropine, an effect m e d i a t e d b y its a c t i o n via m u s c a r i n i c acetylch01ine receptors. A beneficial action of p r a z o s i n has been r e p o r t e d clinically ( K u c k et al., 1981) in left ventricular failure p r e s u m a b l y d u e to a stabilising effect of p r a z o s i n on a l - a d r e n o c e p tots in the m y o c a r d i u m . Studies are u n d e r w a y at p r e s e n t to localise other r e c e p t o r types in the h e a r t a n d to establish whether changes in r e c e p t o r density can b e shown to a c c o m p a n y certain p a t h o l o g i cal conditions.
Acknowledgements We would like to thank DuPont for the gift for naloxone HCI, the British Heart Foundation and Rachel Sykes for excellent assistance.
References Dashwood, M.R., 1983, Central and peripheral prazosin binding: an in vitro autoradiographic study in the rat, European J. Pharmacol. 86, 51. Dashwood, M.R., M.P. Giibey and K.M. Spyer, 1985, The localization of adrenoceptors and opiate receptors in regions of the cat central nervous system involved in cardiovascular control, Neuroscience 15, 537. Dashwood, M.R. and M. Jacobs, 1985, Autoradiographic study of the a-adrenoceptors of rat aorta and tail artery, European J. Pharmacol. 115, 129. Guicheney, P. and P. Meyer, 1981, Binding of [3H]prazosin and [3H]dihydroergocryptine to rat cardiac a-adrenoceptors, Br. J. Pharmacol. 73, 33. Kuck, K.H., P. Hanrath, A. Zenke, D. Mathey and W. Bleifeld, 1981, Haemodynamic effects of long term prazosin therapy in patients with congestive heart failure, J. Cardiovasc. Pharmacol. 2 (Suppl. 3), $427. Nedoma, J., J. Slavikova and S. Tucek, 1986, Muscarinic acetycholine receptors in the hearts of rats before and after birth, Pfliigers Arch. 406, 45. Unnerstall, J.R., T.A. Kopajtic and M.J. Kuhar, 1984, Distribution of a2-agonist binding sites in the rat and human central nervous system: analysis of some functional anatomic correlates of the pharmacologic effects of clonidine and related adrenergic agents, Brain Res. Rev. 7, 69. Wamsley, J.K., M.S. Lewis, W.S. Young and M.J. Kuhar, 1981, Autoradiographic localization of muscarinic .cholinergic receptors in rat brainstem, J. Neurosci. 1, 176. Watanabe, A.M., L.R. Jones, A.S. Manalan and H.R. Besch, Jr., 1982, Cardiac autonomic receptors. Recent concepts from radiolabelled ligand-binding studies, Circ. Res. 50, 161. Young, W.S. and M.J. Kuhar, 1979, A new method for receptor autoradiography: [3H]opioid receptors in rat brain, Brain Res. 179, 255.
279
Elsevier Short communication
AUTORADIOGRAPHIC LOCALIZATION OF a-ADRENOCEPTORS, MUSCARINIC A C E T Y L C H O L I N E R E C E P T O R S AND O P I A T E R E C E P T O R S IN T H E H E A R T MICHAEL R. DASHWOOD * and K. MICHAEL SPYER Department of Physiology, Royal Free Hospital School of Medicine, Rowland Hill Street, London NW3 2PF, England
Received 23 June 1986, accepted 24 June 1986
M.R. DASHWOOD and K.M. SPYER, Autoradiographic localization of a-adrenoceptors, receptors and opiate receptors in the heart, European J. Pharmacol. 127 (1986) 279-282.
muscarinic acetylchofine
Using in vitro autoradiography the distribution of [3H]rauwolscine, [3H]prazosin and [3H]quinuclidinyl benzilate binding sites has been demonstrated in cardiac tissue taken from the cat and rat. A similar distribution of both a 1- and a2-adrenoceptor sites was seen but the distribution of muscarinic acetylcholine sites was markedly different. a-Adrenoceptors were present predominantly in ventricular muscle whereas muscarinic acetylcholine receptors exhibited a greater density in atrial tissue compared to ventricular muscle. Opiate receptors were absent from cardiac tissue. Muscarinic acetylcholine receptors
a-Adrenoceptors
1. Introduction
a-Adrenoceptor, muscarinic acetylcholine receptor and opiate receptor agonists all affect heart rate when administered intravenously. The site of action of these compounds may be directly on the myocardium, mediated centrally or reflexly as a consequence of their effects on arterial blood pressure. Both muscarinic acetylcholine and al-adrenoceptors have been identified in membrane preparations of cardiac muscle (Guicheney and Meyer, 1981; N e d o m a et al., 1986) and eq-adrenoceptors have been shown to exist in the heart using autoradiography (Dashwood, 1983). In the present study we have used the in vitro autoradiographic technique of Young and Kuhar (1979) to localize a 1- and a2-adrenoceptors, muscarinic acetylcholine and opiate receptors in the heart of cat and rat.
* To whom all correspondence should be addressed. 0014-2999/86/$03.50 © 1986 Elsevier Science Publishers B.V.
Ventricular muscle
Autoradiography
Atria
2. Materials and m e t h o d s
Male and female Sprague-Dawley rats (RFHSM-derived, body weight around 250 g) and male and female cats (body weight around 3 kg) were used for this study. Animals were anaesthetized with sodium pentobarbitone (80 rng/kg i.p.) and perfused intracardially with ice-cold 100 m M phosphate buffer containing 0.32 M sucrose (pH 7.4) and lightly fixed with 0.1% formaldehyde solution in phosphate-sucrose buffer. After fixation the heart was removed, rinsed in buffer and mounted in Ames O.C.T. compound onto a cryostat chuck. Serial 20/~m longitudinal sections were taken through the heart at - 2 0 ° C which were thaw-mounted onto gelatinized microscope slides, allowed to dry and stored either overnight at - 2 0 ° C or for up to 4 weeks at - 7 0 ° C . Paired slides were then taken and incubated for 60 min at 4 ° C in 4 nM [3H]naloxone (specific activity 55 C i / m m o l ) made up in 170 m M Tris HC1 buffer containing 100 m M NaC1 (pH 7.4), 60 min at 4 ° C in 5 nM [3H]prazosin (specific activity 33 C i / mmol) made up in 170 m M Tris HC1 buffer (pH
280 7.4) or for 120 min at 4 ° C in 2 nM [3H]rauwolscine (specific activity 80 C i / m m o l ) and 1 nM [3H]quinuclidinyl benzilate (QNB) (specific activity 38 C i / m m o l ) both made up in 100 m M phosphate buffer (pH 7.4) in order to identify opiate, al-adrenoceptors, a2-adrenoceptors and muscarinic acetylcholine receptors respectively. After incubation sections were washed twice in the appropriate buffers before being blown dry in a cold stream of air, placed into 24 x 30 cm X-ray cassettes and apposed to LKB Ultrofilm for 4-6 weeks at 4°C. The degree of non-specific binding to tissue sections was established in the presence of 1/~M naloxone or morphine (opiates), 1-10/~M phentolamine (a~- and a2-adrenoceptors ) and 1 /~M carbachol (muscarinic acetylcholine receptors). After exposure the LKB Ultrofilm was developed in Kodak D19 developer (undiluted) for 5 min at 20°C, fixed in Ilfospeed fixer (diluted 1 to 4 with distilled water) for 5 min at 20°C and rinsed for at least 20 min at 20 °C before drying. Autoradiographs were visually inspected and photographed, where appropriate, using a Nikon Macro system. All tritiated ligands were purchased from Amersham International.
[3H]QNB binding to cardiac muscle in general which represents the presence of muscarinic acetylcholine receptor sites. Seventy three percent of this binding was displaced by incubating sections in the presence of 1 /~M carbachol. There was, however, a regional variation in this binding with the [3H]QNB binding being much greater in ventricular muscle than both a 1- and a2-adrenoceptors. There was a most striking muscarinic acetylcholine receptor presence in the atria (figs. 1A and 2A).
3. Results There were no qualitative differences in the distribution of the various receptors in the heart of both cats and rats. Opiate receptors, indicated by [3H]naloxone binding, were absent from all regions of the heart (figs. 1B and 2B). Both cqand a2-adrenoceptors ([3H]prazosin and [3H]rauwolscine binding) exhibited a fairly high and variable distribution over ventricular muscle with a lower degree of binding to atrial tissue (figs. 1C-D and 2C-D). This binding was to specific a 1- and a2-adrenoceptor sites since both [3H]prazosin and [3H]rauwolscine were displaced to a marked degree (about 80%) in those sections incubated in the presence of 1/~M unlabelled phentolamine. It is interesting to note that there was a high degree of [ 3H]prazosin binding to the inner surface of the aorta (arrowed in fig. 2C) whereas there was no [3H]rauwolscine binding in the adjacent section (fig. 2D). There was an extremely high degree of
Fig. 1. Autoradiographic localization of receptors in consecutive sections of rat heart. (A) Section incubated in 1 nM [3H]QNB. There is a very dense binding at the atria (A) with a lower distribution over the ventricular muscle (V). (B) Section incubated in 4 nM [3H]naloxone. There is no [3H]naloxone binding to cardiac tissue. (C) Section incubated in 5 nM [3H]prazosin. There is a generalized moderate distribution of binding over the ventricular muscle with a reduced binding at atrial regions. (D) Section incubated in 2 nM [3H]rauwolscine. There is a generalized moderate distribution of binding over the ventricular muscle with a slightly reduced binding at atrial regions. Scale bar = 2.5 mm.
281
Fig. 2. Autoradiographic localization of receptors in consecutive sections of cat heart. (A) Section incubated in 1 nM [3H]QNB. There is a very dense binding to ventricular muscle (V) accompanied by extremely dense binding at atrial areas (A). (B) Section incubated in 4 nM [3H]naloxone. There is no binding to cardiac tissue. (C) Section incubated in 5 nM [3H]prazosin. There is a generalized high binding to ventricular muscle with a much weaker binding to the atria. There is a strong binding of [3H]prazosin to the internal surface of the aorta (arrowed). (D) Section incubated in 2 mM [3H]rauwolscine. There is a generalized high binding to ventricular muscle with a weaker binding to the atria. Scale bar = 5 mm.
4. Discussion O p i a t e receptor, muscarinic acetylcholine receptor and ct-adrenoceptor agonists all elicit effects on the heart when administered intravenously. These effects m a y be mediated centrally since receptors for these c o m p o u n d s have been identified in brain areas associated with cardiovascular control (Young and Kuhar, 1979; W a m s ley et al., 1981; Unnerstall et al., 1984).
The purpose of the present autoradiographic study was to establish whether opiate receptors, muscarinic acetylcholine receptors and a-adrenoceptors exist in the heart of the cat and rat where direct a c t i o n s m a y be elicited. There was no [3H]naloxone binding to cardiac tissue of either species (figs. 1B and 2B). Heart sections were incubated in parallel with brain and spinal cord tissue which exhibited areas of high [3H]naloxone binding in, for example, the nucleus of the tractus solitarii and substantia gelatinosa of the thoracic spinal cord ( D a s h w o o d et ak, 1985). This result would indicate that opiate receptors are absent from cardiac tissue and that opiate-induced bradycardia may be due, in part, to an effect on receptors within the dorsomedial brainstem (see D a s h w o o d et al., 1985). There was a fairly dense distribution of both a 1 ([3H]prazosin binding)and a 2 ([3H]rauwolscine binding)-adrenoceptors on the heart. The distribution was similar in tissue from both cat and rat (figs. 1 and 2). The highest density of a-adrenoceptors was in ventricular muscle with a reduced binding at atrial regions (figs. 1C-D and 2C-D). Subtle variations in receptor density m a y well have been present across sections of ventricular tissue. There certainly appears to be a higher degree of [3H]QNB binding at the epicardium than endocardium. However there was no obvious increase in any receptor density at specialized areas of the heart such as the SA or A V node. Subsequent densitometric analysis of these autoradiographs m a y reveal subtle regional variations in receptor distribution. Figure 2 A - D illustrate four consecutive 20 /~m sections through the heart. It is interesting to note that in this particular plane a strong binding of [3H]prazosin is evident on the internal surface of the aorta whereas [3H]rauwolscine binding is absent (compare fig. 2C and D). This result is in agreement with the recent data of D a s h w o o d and Jacobs (1985) who showed a presence of al-adrenoceptors on the rat aorta but an absence of a 2adrenoceptors. Although there is considerable evidence for the presence of a~-adrenoceptors in cardiac muscle (Guicheney and Meyer, 1981), there is little substantial evidence for an a2-adrenoce ptor presence. According to the review of W a t a n a b e et al. (1982) the majority of ligand binding studies
282 carried out in c a r d i a c muscle have been with [3H]dihydroergocryptine, an a - a d r e n o c e p t o r antagonist, which will n o t d i s c r i m i n a t e between a 1a n d a2-subtypes. T h e most striking b i n d i n g o b s e r v e d in this s t u d y was that of [ 3 H ] Q N B to c a r d i a c tissue. T h e b i n d ing to ventricular muscle was stronger than that of [3H]prazosin a n d [3H]rauwolscine ( c o m p a r e figs. 1A a n d 2 A with figs. 1 C - D a n d 2 C - D ) but, whereas a - a d r e n o c e p t o r s were a l m o s t a b s e n t from atrial regions there was an e x t r e m e l y dense m u s c a r i n i c acetylcholine r e c e p t o r presence at this level (figs. 1A a n d 2A). These results not o n l y c o n f i r m results from r e c e p t o r b i n d i n g studies using h o m o g e n a t e prep a r a t i o n s of c a r d i a c tissue for a l - a d r e n o c e p t o r s a n d m u s c a r i n i c acetylcholine receptors (Guicheney a n d Meyer, 1981; N e d o m a et al., 1986) b u t also give an i n d i c a t i o n of the gross differential d i s t r i b u t i o n of such receptors across the heart. T h e r e are some i n d i c a t i o n s of the functional significance of these receptors. It is recognized that vagal efferent s t i m u l a t i o n causes a p r o f o u n d b r a d y c a r d i a which is b l o c k e d b y atropine, an effect m e d i a t e d b y its a c t i o n via m u s c a r i n i c acetylch01ine receptors. A beneficial action of p r a z o s i n has been r e p o r t e d clinically ( K u c k et al., 1981) in left ventricular failure p r e s u m a b l y d u e to a stabilising effect of p r a z o s i n on a l - a d r e n o c e p tots in the m y o c a r d i u m . Studies are u n d e r w a y at p r e s e n t to localise other r e c e p t o r types in the h e a r t a n d to establish whether changes in r e c e p t o r density can b e shown to a c c o m p a n y certain p a t h o l o g i cal conditions.
Acknowledgements We would like to thank DuPont for the gift for naloxone HCI, the British Heart Foundation and Rachel Sykes for excellent assistance.
References Dashwood, M.R., 1983, Central and peripheral prazosin binding: an in vitro autoradiographic study in the rat, European J. Pharmacol. 86, 51. Dashwood, M.R., M.P. Giibey and K.M. Spyer, 1985, The localization of adrenoceptors and opiate receptors in regions of the cat central nervous system involved in cardiovascular control, Neuroscience 15, 537. Dashwood, M.R. and M. Jacobs, 1985, Autoradiographic study of the a-adrenoceptors of rat aorta and tail artery, European J. Pharmacol. 115, 129. Guicheney, P. and P. Meyer, 1981, Binding of [3H]prazosin and [3H]dihydroergocryptine to rat cardiac a-adrenoceptors, Br. J. Pharmacol. 73, 33. Kuck, K.H., P. Hanrath, A. Zenke, D. Mathey and W. Bleifeld, 1981, Haemodynamic effects of long term prazosin therapy in patients with congestive heart failure, J. Cardiovasc. Pharmacol. 2 (Suppl. 3), $427. Nedoma, J., J. Slavikova and S. Tucek, 1986, Muscarinic acetycholine receptors in the hearts of rats before and after birth, Pfliigers Arch. 406, 45. Unnerstall, J.R., T.A. Kopajtic and M.J. Kuhar, 1984, Distribution of a2-agonist binding sites in the rat and human central nervous system: analysis of some functional anatomic correlates of the pharmacologic effects of clonidine and related adrenergic agents, Brain Res. Rev. 7, 69. Wamsley, J.K., M.S. Lewis, W.S. Young and M.J. Kuhar, 1981, Autoradiographic localization of muscarinic .cholinergic receptors in rat brainstem, J. Neurosci. 1, 176. Watanabe, A.M., L.R. Jones, A.S. Manalan and H.R. Besch, Jr., 1982, Cardiac autonomic receptors. Recent concepts from radiolabelled ligand-binding studies, Circ. Res. 50, 161. Young, W.S. and M.J. Kuhar, 1979, A new method for receptor autoradiography: [3H]opioid receptors in rat brain, Brain Res. 179, 255.