Pharmacology 18: 72-79 (1979)
Effects of Urotensin I on the Isolated Rat Tail Artery1 M E . G erritsen and K. Lederis2 Division o f Pharmacology and Therapeutics, Faculty o f Medicine, University o f Calgary, Calgary, Alta.
Key Words. Rat tail artery • Vasodilation • Hypotensive peptides and urotensin Abstract. Urotensin I (UI) was found to elicit dose-related relaxation responses in isolated
helical strips o f tire rat tail artery. The responses were not prevented by adrenergic, cholingergic or histaminergic blocking agents. Competitive and non-competitive components o f antagonism were observed to noradrenaline-, 5-hydroxytryptamine-, and arginine vasopressin-induced contractions. Atropine caused a direct relaxation o f the isolated vascular tissues, as well as a significant potentiation o f UI responses.
Urotensin 1 (U I), one o f two peptides iso lated from the urophysis o f the white sucker
venous administration o f low doses o f UI in the
(Catastomus commersoni) (19) has been shown
pronounced vasodilatation in the mesenteric
to have a long lasting hypotensive action in a
region
number o f mammalian species (13, 15).
vascular beds (15).
Mecamylamine, hexamethonium bromide,
anaesthetized dog were found to produce a without noticeable effects
in other
In the present study, helical strips o f isolated
propranolol and diphenhydramine did not alter
rat tail arteries were used, firstly, to evaluate
the blood pressure lowering effect o f UI in
this tissue as an assay preparation for U I, and
conscious
secondly, to elucidate the mode o f action o f
rats
or
tire
perfusion
pressure
lowering effect in the isolated rat hind limb
this peptide in vascular tissue.
(13, 14) suggesting that UI elicited hypotension in the rat by a direct dilatory action in the resistance vessels, accompanied by a reflex
Methods
1 Supported by M R C (Canada). 2 M .E .G is a recipient o f M RC Studentship, Izaac Walton Killam Merit Award and K .L. is an M R C Associate.
Male Sprague-Dawley rats (200-400 g body weight) were decapitated and the ventral tail artery removed. Helical strips 1 - 2 X 1 0 -1 2 mm were pre pared and suspended at 500 mg o f tension in 7-ml tissue baths perfused with a Krebs-Henseleit buffer at 37 °C, aerated with 95% 0 2, 5% C 0 2. The vascular
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tachycardia. Close-arterial or systemic intra
Urotensin I Effects on Rat Tail Artery
The following drugs were used: noradrenaline (Levophed bitartrate, Winthrop), 5-hydroxytryptamine (Sigma), phenoxybenzamine (PBZ: Sm ith, Kline, French), phentolamine (Rogitine, Ciba), acetylcholine chloride (Sigma), histamine dihydrochloride (BDH), mepyramine maleate (Poulenc), metiamide (Smith, Kline, French), burimamide (Smith, Kline, French), potassium chloride (reagent grade, Fisher) and arginine vasopressin (A VP; Pitressin, Parkc-Davis). All drugs, with the exception o f U l, were freshly prepared and made up in concentrated stock solutions and then diluted to appropriate concentrations in 0.9% NaCl. The volume added to the 7-ml organ baths never exceeded 0.1 ml. PBZ was made up in a concentrated acid-ethanol solution. Tissues were incubated in the
Fig. 1. Comparative log-dose response curves for UI in the isolat ed rat tail artery given elevated vas cular tone by 8.4 x 10"’ M NA (•), 10' ' M 5-HT (▼ ), or 23 nvW KC1 (o).
presence o f PBZ for 20 min prior to the experiment. Partially purified UI in 0.1 N HC1 was kindly supplied by Dr. Denis McMaster. The activity o f U l is expressed in units as defined by Bern and Lederis (1). Variations in purity o f urotensin preparations and subsequent varia tions in bioassay account for discrepancies in position of log-dose response curves from one group o f experiments to the next. However, all groups compared utilized a single purified lot o f UI. Responses to drugs were expressed as percentage change from the equilibrium tension attained by N A , 5-HT or KC1. Am ounts o f drugs stated refer to the salt/ml bath fluid. Ion concentrations were determined by volumetric additions o f gravimctrically determined stock solutions.
Results
Responsesof the Isolated Rat Tail Artery to
UI In different experiments, either N A , 5-HT or KC1 were used to elevate the tension o f vascular muscle for subsequent administration o f U I.
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strips were allowed to equilibrate for 90 min prior to starting experiments. Isometric responses to drugs were detected by means o f a Statham force-displace ment transducer (No. UC-2) and recorded on a Beck man dynograph. To allow observation o f relaxation responses, vascular tone was elevated by the inclusion in the perfusion mixture o f either 23 mM KC1, 8.4 X 1 0 '’ M noradrenaline (N A), or 1.4 X 10"’ M 5-hydrotryptamine (5-HT).
73
74
Gerritsen/Lederis
Cumulative dose-response curves for U1 were obtained for rat tail arteries using each o f the three contractile agents; the results are sum marized in figure 1. Minimum effective doses o f UI with all three contractile agents were o f the order o f 1 mU/ml. Maximum effective doses
Table I. Time o f onset and percent relaxation (x ± SEM . n = 6) for two doses o f UI (1 = 7.6 mU/ml UI, 2 = 15.2mU/ml UI) in preparations given elevated vascular tone by N A , 5-HT or KCl
Contractile agent
Percent relaxation
Time o f onset, sec
NA
1 2 1 2 1 2
78.6 52.5 37.1 13.2 9.1 4.9
were not determined as purified extracts current ly available were not sufficiently high in activity, thus requiring large volumes (in excess o f 0.1 ml o f extract) to attain doses greater than 125 mU/ ml. Maximum responses to UI were attained
5-HT KCl
within three minutes and responses were o f long
9.0 30.2 9.6 30.8 39.5 79.5
± 0.9
t 2.8 t 1.0 ± 1.2 ± 4.5* ± 9.5*
± ± t ± ± ±
3.2 6.1 9.0* 3.9* 2.0* 1.1*
duration (10 min to several hours) making a detailed study o f dose-duration relationships difficult.
* p
Effects of Urotensin I on the Isolated Rat Tail Artery1 M E . G erritsen and K. Lederis2 Division o f Pharmacology and Therapeutics, Faculty o f Medicine, University o f Calgary, Calgary, Alta.
Key Words. Rat tail artery • Vasodilation • Hypotensive peptides and urotensin Abstract. Urotensin I (UI) was found to elicit dose-related relaxation responses in isolated
helical strips o f tire rat tail artery. The responses were not prevented by adrenergic, cholingergic or histaminergic blocking agents. Competitive and non-competitive components o f antagonism were observed to noradrenaline-, 5-hydroxytryptamine-, and arginine vasopressin-induced contractions. Atropine caused a direct relaxation o f the isolated vascular tissues, as well as a significant potentiation o f UI responses.
Urotensin 1 (U I), one o f two peptides iso lated from the urophysis o f the white sucker
venous administration o f low doses o f UI in the
(Catastomus commersoni) (19) has been shown
pronounced vasodilatation in the mesenteric
to have a long lasting hypotensive action in a
region
number o f mammalian species (13, 15).
vascular beds (15).
Mecamylamine, hexamethonium bromide,
anaesthetized dog were found to produce a without noticeable effects
in other
In the present study, helical strips o f isolated
propranolol and diphenhydramine did not alter
rat tail arteries were used, firstly, to evaluate
the blood pressure lowering effect o f UI in
this tissue as an assay preparation for U I, and
conscious
secondly, to elucidate the mode o f action o f
rats
or
tire
perfusion
pressure
lowering effect in the isolated rat hind limb
this peptide in vascular tissue.
(13, 14) suggesting that UI elicited hypotension in the rat by a direct dilatory action in the resistance vessels, accompanied by a reflex
Methods
1 Supported by M R C (Canada). 2 M .E .G is a recipient o f M RC Studentship, Izaac Walton Killam Merit Award and K .L. is an M R C Associate.
Male Sprague-Dawley rats (200-400 g body weight) were decapitated and the ventral tail artery removed. Helical strips 1 - 2 X 1 0 -1 2 mm were pre pared and suspended at 500 mg o f tension in 7-ml tissue baths perfused with a Krebs-Henseleit buffer at 37 °C, aerated with 95% 0 2, 5% C 0 2. The vascular
Downloaded by: Lund University Libraries 130.235.66.10 - 1/1/2019 7:15:30 PM
tachycardia. Close-arterial or systemic intra
Urotensin I Effects on Rat Tail Artery
The following drugs were used: noradrenaline (Levophed bitartrate, Winthrop), 5-hydroxytryptamine (Sigma), phenoxybenzamine (PBZ: Sm ith, Kline, French), phentolamine (Rogitine, Ciba), acetylcholine chloride (Sigma), histamine dihydrochloride (BDH), mepyramine maleate (Poulenc), metiamide (Smith, Kline, French), burimamide (Smith, Kline, French), potassium chloride (reagent grade, Fisher) and arginine vasopressin (A VP; Pitressin, Parkc-Davis). All drugs, with the exception o f U l, were freshly prepared and made up in concentrated stock solutions and then diluted to appropriate concentrations in 0.9% NaCl. The volume added to the 7-ml organ baths never exceeded 0.1 ml. PBZ was made up in a concentrated acid-ethanol solution. Tissues were incubated in the
Fig. 1. Comparative log-dose response curves for UI in the isolat ed rat tail artery given elevated vas cular tone by 8.4 x 10"’ M NA (•), 10' ' M 5-HT (▼ ), or 23 nvW KC1 (o).
presence o f PBZ for 20 min prior to the experiment. Partially purified UI in 0.1 N HC1 was kindly supplied by Dr. Denis McMaster. The activity o f U l is expressed in units as defined by Bern and Lederis (1). Variations in purity o f urotensin preparations and subsequent varia tions in bioassay account for discrepancies in position of log-dose response curves from one group o f experiments to the next. However, all groups compared utilized a single purified lot o f UI. Responses to drugs were expressed as percentage change from the equilibrium tension attained by N A , 5-HT or KC1. Am ounts o f drugs stated refer to the salt/ml bath fluid. Ion concentrations were determined by volumetric additions o f gravimctrically determined stock solutions.
Results
Responsesof the Isolated Rat Tail Artery to
UI In different experiments, either N A , 5-HT or KC1 were used to elevate the tension o f vascular muscle for subsequent administration o f U I.
Downloaded by: Lund University Libraries 130.235.66.10 - 1/1/2019 7:15:30 PM
strips were allowed to equilibrate for 90 min prior to starting experiments. Isometric responses to drugs were detected by means o f a Statham force-displace ment transducer (No. UC-2) and recorded on a Beck man dynograph. To allow observation o f relaxation responses, vascular tone was elevated by the inclusion in the perfusion mixture o f either 23 mM KC1, 8.4 X 1 0 '’ M noradrenaline (N A), or 1.4 X 10"’ M 5-hydrotryptamine (5-HT).
73
74
Gerritsen/Lederis
Cumulative dose-response curves for U1 were obtained for rat tail arteries using each o f the three contractile agents; the results are sum marized in figure 1. Minimum effective doses o f UI with all three contractile agents were o f the order o f 1 mU/ml. Maximum effective doses
Table I. Time o f onset and percent relaxation (x ± SEM . n = 6) for two doses o f UI (1 = 7.6 mU/ml UI, 2 = 15.2mU/ml UI) in preparations given elevated vascular tone by N A , 5-HT or KCl
Contractile agent
Percent relaxation
Time o f onset, sec
NA
1 2 1 2 1 2
78.6 52.5 37.1 13.2 9.1 4.9
were not determined as purified extracts current ly available were not sufficiently high in activity, thus requiring large volumes (in excess o f 0.1 ml o f extract) to attain doses greater than 125 mU/ ml. Maximum responses to UI were attained
5-HT KCl
within three minutes and responses were o f long
9.0 30.2 9.6 30.8 39.5 79.5
± 0.9
t 2.8 t 1.0 ± 1.2 ± 4.5* ± 9.5*
± ± t ± ± ±
3.2 6.1 9.0* 3.9* 2.0* 1.1*
duration (10 min to several hours) making a detailed study o f dose-duration relationships difficult.
* p
