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Central cardiovascular effects of AVP and ANP in normotensive and spontaneously hypertensive rats
Journal of the Autonomic Nervous System, 47 (1994) 33-43
33
© 1994 Elsevier Science B.V. All rights reserved 0165-1838/94/$07.00 JANS 01480
Central cardiovascular effects of AVP and ANP in normotensive and spontaneously hypertensive rats Konrad St~pniakowski a, Adam Budzikowski b, S/awomir Loft b and Ewa Szczepafiska-Sadowska b,, a Departments of Hypertension and Angiology and b Department of Clinical and Applied Physiology, MedicalAcademy of Warsaw, 00-927 Warsaw, Poland (Received 14 April 1993) (Revision received 20 June 1993) (Accepted 12 July 1993)
Key words: Vasopressin; Atrial natriuretic peptide; Blood pressure; Central actions; SHR; WKY Abstract The purpose of the present study was to compare influence of central arginine vasopressin (AVP) and of atrial natriuretic peptide (ANP) on control of arterial blood pressure (MAP) and heart rate (HR) in normotensive (WKY) and spontaneously hypertensive (SHR) rats. Three series of experiments were performed on 30 WKY and 30 SHR, chronically instrumented with guide tubes in the lateral ventricle (LV) and arterial and venous catheters. MAP and HR were monitored before and after i.v. injections of either vehicle or 1, 10 and 50 ng of AVP and 25, 125 and 500 ng of ANP. Sensitivity of cardiac component of baroreflex (CCB), expressed as a slope of the regression line was determined from relationships between systolic arterial pressure (SAP) and HR period (HRp) during phenylephrine (Phe)-induced hypertension and sodium nitroprusside (SN)-induced hypotension. CCB was measured before and after administration of either vehicle, AVP, ANP, or both peptides together. Increases of MAP occurred after LV administration of 1, 10 and 50 ng of AVP in WKY and of 10 and 50 ng in SHR. ANP did not cause significant changes in MAP in both strains as compared to vehicle, but it abolished AVP-induced MAP increase in WKY and SHR. CCB was reduced in WKY and SHR after LV administration of AVP during SN-induced hypotension. In SHR but not in WKY administration of ANP, AVP and ANP + AVP decreased CCB during Phe-induced MAP elevation. The results indicate that centrally applied AVP and ANP exert differential effects on blood pressure and baroreflex control of heart rate in WKY and SHR and suggest interaction of these two peptides in blood pressure regulation at the level of central nervous system.
Introduction
Many recent investigations suggest the existence of mutual interactions between atrial natriuretic peptide (ANP) and vasopressin (AVP) in regulation of body fluid balance and blood pressure. Systemically administered ANP has been
* Corresponding author. Department of Clinical and Applied Physiology, Medical Academy of Warsaw, Krakowskie Przedmiescie 26/28 00-927 Warsaw, Poland. Tel.: 26-45-86.
SSDI 0 1 6 5 - 1 8 3 8 ( 9 3 ) E 0 0 9 7 - O
found to reduce blood pressure increase and to enhance reflex bradycardia elicited by systemic vasopressin [16,38]. Immunohistochemical evidence indicates the similar location of ANP and AVP and their receptors in forebrain and brain stem areas suggesting that these peptides contribute to blood pressure regulation [4,10,17,18,28,29,39]. Significant differences in AVP content and density of ANP binding sites in spontaneously hypertensive (SHR) and normotensive (WKY) rats [20,21,25,27,34,35] suggest the role of these two neuropeptides in the
34
pathogenesis of hypertension. The purpose of the present study was to compare blood pressure and heart rate responses to central administration of AVP and ANP as well as the interaction between these two peptides in blood pressure and heart rate regulation in normotensive (WKY) and spontaneously hypertensive (SHR) rats. We have found that central effects of vasopressin and ANP on blood pressure and the sensitivity of cardiac component of baroreflex (CCB) markedly differ in WKY and SHR. Moreover, our results indicate that ANP suppresses AVP-induced pressor responses in WKY and SHR and modifies AVP-induced changes in sensitivity of the cardiac component of baroreflex in WKY.
Material and Methods
Material Experiments were performed on conscious, freely moving SHR and WKY, males, 12-14 weeks old (body weight between 280-300 g), maintained on a 12-h l i g h t / d a r k cycle (light from 07.00 to 19.00 h). Before the experiments they had free access to water and commercial laboratory chow, containing 0.5% NaCI. Food and water were removed during the experimental sessions.
Surgical procedures Seven to 10 days before the experiment the animals were anesthetized with chloral hydrate (chloral-hydrate, Merck, 36 mg/100g i.p.). A stainless steel guide tube (O.D. 0.6 mm) was implanted into the lateral cerebral ventricle (LV) through the opening drilled in the skull according to the following parameters: - 1 . 5 mm posterior to the bregma, 2 mm lateral to the sagittal suture and 5 mm below the surface of the skull. The cannula was closed with a stainless steel stylet and fixed in the skull with acrylic cement. One or 2 days prior to the experiment all rats were subjected to ether anesthesia and instrumented with arterial catheter (PE 20) for blood pressure measurements during freely moving state. The catheter was inserted into the aorta through the left femoral artery. In the third group of animals
the additional PE 20 catheter was introduced into the vena cava through the right femoral vein for injections of phenylephrine (Phe) and sodium nitroprusside (SN). The catheters were connected to PE 60 tubing filled with saline and heparin, plugged with stoppers, tunnelled under the skin and exteriorized on the neck. After the surgery the animals were placed in individual experimental cages for recovery.
Experimental protocol The experiments were performed on 30 WKY and 30 SHR having icv cannulae appropriately located in the LV. The animals were divided into three experimental groups, each consisting of t0 WKY and 10 SHR. Mean values of resting MAP and heart rate (HR) found in the three experimental groups are presented in Tables I and II1. At the beginning of the experiment the arterial catheter was connected to the blood pressure unit for MAP and H R measurements. The stylet was removed from the LV cannula. The actual experiments started after 30-40 min of rest to allow for stabilization of MAP and HR. LV injections were made by means of Hamilton microsyringe at a rate of 5 / z l / 1 0 s. In Series 1 effect of central AVP on MAP and H R was compared in 10 WKY and 10 SHR. At the beginning of each experimental session the animals received LV injection of vehicle (5 pA). This was followed by administration of 1, 10 and 50 ng of AVP in 5 ~1 of vehicle. Subsequent injections were performed with time intervals of at least 40 min when MAP and H R returned to the resting level (Table I). Blood pressure -systolic (SAP), diastolic (DAP) and heart rate were monitored every 1 min during 10 min and every 5 rain during subsequent 30 min following the injection. To assess the effect of central AVP on the relationship between systolic arterial pressure (SAP) and heart rate period (HRp) individual values of SAP and corresponding (beat-to-beat) HRp during the first 10 min after icv administration of either vehicle or AVP were subjected to regression analysis. Slopes of the regression lines significant at the level of P < 0.001 were subjected to A N O V A to estimate significance of differences between treatments and strains.
35 TABLE I Mean blood pressure and heart rate calues preceding L V AVP and ANP injections
Series 1
Vehicle 1 ngAVP
MAP (mmHg) 114_+ 3 113_+ 3 HR (b/ rain) 327_+11 339_+12
10ngAVP
50ngAVP
117_+ 3
117_+ 3
339_+12
351_+ 9
176_+ 3
1795:4
406_+ 1
426_+13
SI-IR
MAP (mmHg) 178_+ 2 175-+ 3 HR (b/ rain) 407-+12 396_+12
Series 2 ACSF 10 ng ANP 125 ng ANP 500 ng ANP WKY MAP (mmHg) 97J: 4 99+ 3 94-+ 3 96+ 3 HR (b/ min) 321+ 8 339+ 9 317_+ 9 334+ 4 SItR
MAP (mmHg) 185_+ 4 185+ 4 HR (b/ min) 402+10 386+12
185_+ 3
188+ 3
4025:11
406+12
Values are as mean+SEM. ACSF, artificial cerebrospinal fluid; AVP, arginine vasopressin, ANP, atrial natriuretic peptide; WKY, normotensive Wistar Kyoto rats; SHR, spontaneously hypertensive rats. Effect of central administration of A N P on M A P and H R was compared in Series 2. The animals received LV injections of either vehicle or with 25, 125 and 500 ng of atrial natriuretic peptide (ANP) in 5/zl of vehicle. The experimental procedure was identical as in Series 1. The relationships between SAP and H R p during the first 10 min after administration of either artificial cerebrospinal fluid (ACSF) or A N P were determined and subjected to regression analysis as in Series 1. Series 3 was conducted to find out whether central A N P and A V P interact in blood pressure regulation. The animals were subjected to LV administrations of (1) vehicle, (2) 125 ng of ANP, (3) 10 ng of AVP and 4) 10 ng AVP + 125 ng ANP, at the minimum of at least 40-min intervals. Ten minutes after each LV injection phenyl-
ephrine (Phe, 0.5 /zg/100 g) was given i.v. as a bolus in 100 /~1 of 0.9% NaCI and its effects on blood pressure and heart rate recorded continuously. When M A P and H R stabilized to the level found prior to Phe administration (usually 5 - 7 min after i.v. Phe injection) the rats were injected i.v. with sodium nitroprusside (SN, 3 /~g/100 g) in 100 /3,1 0.9% NaC1, while blood pressure and heart rate were recorded continuously. After completion of the experiments the position of the cannula in the lateral ventricle was verified by injection of Evans Blue and postm o r t e m brain examination. In some animals the appropriate location of the cannula was preliminary verified by testing presence of drinking and pressor response to LV administration of 100 ng of angiotensin II. Measurements
For blood pressure and heart rate measurements the arterial catheter was connected to the blood pressure recording device consisting of transducer and amplifier (Statham, Gould P 23Db). The signal frequency was modulated with analog to digital converter connected through an interface to Amstrad-Schneider CPC 6128 computer which calculated on line the following parameters: systolic (SAP, mmHg), diastolic (DAP, m m H g ) mean (MAP, m m H g ) pressure and heart rate period (HRp, ms). The latter corresponded to the interval between consecutive SAP. To assess sensitivity of the cardiac component of the baroreflex (CCB) [35] individual regression analysis were performed for relationships between SAP and the following H R p from the onset till the maximum of the pressor and depressor responses to Phe and SN, respectively [8]. Changes in CCB after each treatments were evaluated on the basis of differences in slopes of significant regressions. To this end, individual regression coefficients, significant at a level of no less than P < 0.01 were subjected to factorial A N O V A , with treatments and strains as factors. Drugs
Arginine vasopressin (arginine-8-vasopressin, Calbiochem) was kept frozen as a stock solution in 0.1 M acetic acid and freshly diluted with
3~)
artificial cerebrospinal fluid (ACSF) to the required concentration. Appropriate dilution of 0.1 M acetic acid with ACSF served as vehicle. Atrial natriuretic peptide (rat-28 ANF, Peninsula) was kept lyophilized and freshly diluted with ACSF to the required concentration. The ACSF served as vehicle in this series of experiments. No significant difference was noted in heart rate and pressor responses in WKY and SHR to control injections of the vehicles used in Series 1, 2 and 3. The ACSF contained (mM) Na + - 156.0; K + 3.0; Ca 2 + - 1.5; CI- 147.0; H C O ~ 15.
A --,s---
vehicle
+
!
ng
AVP
15i
x
x x ×
× x ×
×x×
x x
x ×
× x 5.
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× x x
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5
ol
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. . . .
5
. . . .
10 . . . .
t 1 1 ~ 1 1 1 1 1 1 1 1 1 k l l ~
15 . . . .
20
. . . .
25
. . . .
I I I I I 1 ~
30
. . . .
35
. . . .
J 40
Results
Central effects of vasopressm and of atrial natriuretic peptide Mean values of MAP and H R preceding injections of AVP and ANP in WKY and SHR are presented in Table I. Administration of 1, 10 and 50 ng of AVP in WKY and of 10 and 50 ng in SHR elicited significant increases of blood pressure (Figs. 1 and 2). In WKY MAP responses
45
rnin
B
Statistical analysis Means and standard errors are presented in the study. Student's t-test, single factor and factorial analysis of variance (ANOVA) for repeated measurements were applied to appropriate data. In each series consecutive values of MAP and HRp after LV injections were compared to the baseline pre-injection values by single factor A N O V A for repeated measurements. The differences in changes in MAP and H R p between various treatments or between WKY and SHR were evaluated by factorial A N O V A on differences from the preinjection level. The NewmanKeul's a posteriori test or multiple comparisons were used to isolate significant differences between individual means [32]. Changes in MAP and H R p elicited by various treatments at a given time were compared by t-test. Linear analysis of regression was performed by the least squares method. The slopes of the regression lines between groups were compared with ANOVA. The results were considered to be significant if P < 0.05.
~1
. . . .
--e--
ng
10
vehicle
AVP 15
10
x x x
× Xx×
×
×
ol
EL
<
-5
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. . . .
5
. . . .
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IIIII 15 . . . .
20
. . . .
IIII 25
. . . .
III 30
II
. . . .
llllllllIl 35
. . . .
40
. . . .
45
c --e.--
vehicle
+
ng
50
AVP 15 x lO
×
x xX xX x
xx
×
~
×
Z
5 v
°
<:]
-5
--10
I I I ; l l l l l L I I I I I l l l
• 0 ....
5 ....
10 ....
J
15 ....
I l l l l l i l l l l l i
20 ....
25 ....
I L I I I I l i l J l _
30 ....
35.
,40 ....
45 rain
Fig. 1. Mean arterial blood pressure changes after LV injection of vehicle (empty circles) and 1 (A), 10 (B) and 50 (C) ng of A V P (solid circles) in WKY. * Significant difference in A V P vs. vehicle changes; (* P < 0.05, * * P < 0.01, * * * P < 0.001). x Significant difference in comparison to initial value; (x p < 0.05, x× p < 0.01, x x x p < 0.001).
after 10 and 50 ng of AVP were biphasic with peaks at 10 and 30-35 min whereas in SHR the same doses resulted in monophasic prolonged increase of blood pressure. In SHR the lowest dose of AVP elicited small decrease in MAP
37
A --e--
vehicle
--o--
1
in HRp (Fig. 3). Changes of HRp after injection o f 50 ng o f A V P in W K Y and S H R w e r e significantly different from each other by two-way A N O V A I F ( l , 1 8 ) = 7.03; P < 0.05].
ng
AVP
In both strains central administration of 10
10
and 50 ng o f A V P markedly r e d u c e d the slopes o f
5. <]
A
-
lO .0 ....
5 ....
t0 ....
15 . . . .
20 ....
25 ....
30 ....
35 ....
40 ....
.-e-- vehicle
45
+
1 ng AVP
rain
B --e-vehicle
---e--
10
15 ~ i
ng
9
3
AVP 15
x 10
x
x
-3
x
x -g
[
5 -15
Q" <
i , i l l l l l l l l l l l l l l l l l l l l l l l l ' l l l l l l l l t l l l l l ~ l • 0 ....
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15 . . . .
20 ....
25 ....
30 ....
35 ....
40 ....
45
rain -5
B
-10 .0 ....
5 ....
t0 ....
15 . . . .
20 ....
25 ....
30 ....
35 ....
40 ....
45
-.-e-- 1 0
--e-- vehicle
min
ng
AVP
C --e-- vehicle
--e-- 5 0
no
AVP
xx x xX x xx x _
~
10
x x x
x x ~
x x x
x x x
x x
x
x
E x x -g xx _1~
x
x
l l l l l l l l l l J l l l l l l l l l l l l l l l l l l l l l l l l t l l l X ~ l l l .0...5
.....
t0 ....
15 . . . .
20 ....
25 ....
30 ....
35 ....
40 ....
45
min <]
-5 -10 •
0
.
.
.
.
5
.
.
.
.
10 .
.
.
.
15 .
.
.
.
20
.
.
.
.
25
.
.
.
.
30
.
.
.
.
35
.
.
.
.
4 0
.
.
.
.
C
45
rain --e-- vehicle
Fig. 2. Mean arterial blood pressure changes after LV injection of vehicle and 1 (A), 10 (B) and 50 (C) ng of AVP in SHR. For other explanations see Fig. 1.
by two-way A N O V A [ F ( 1 , 1 8 ) = 7.17; P < 0.05]. In W K Y H R was not significantly affected,
whereas in SHR injection of 10 and 50 ng of AVP increased HR as shown by significant decreases
ng
AVP
~
which, however, was not different from that observed after LV vehicle in this strain. Changes of MAP after the lowest dose of AVP in WKY and SHR were significantly different from each other
--e-- 5 0
3
- g
-15. o :' i i '51' i iJ i' ','51'' '2'0'." '2'51'. i i;'ii '3'511'i~:ii :,5 rain
Fig. 3. Heart rate period changes after LV injection of 1 (A), 10 (B) and 50 (12) ng of AVP in SHR in comparison to control LV injection of vehicle. For other explanations see Fig. 1.
TABLE II
" F A B L E III
Slopes of the regression lines describing relationship between SAP and HRp under control conditions (Vehich,) and after administration ~# A VP and ANP
Mean blood pressure and heart rate t'alues be~ibre L V inlecttons of A VP and A N P in Series 3 ACSF
Treatment
WKY
SHR
Vehicle AVP I ng 10ng 50ng Vehicle ANP 25 ng 125 ng 5110 ng
1.26
0.71
< 0.001
1.24 1.16 * 1.1 * * * 1.39
0.72 0.65 ** 0.59 * * * 0.67
< 1t.001 <0.001 <0.00t < 0.001
1.35 1.42 1.34
0.66 0.65 0.62 *
P (WKY vs. SHR)
< 0.00l < 0.001 < 0.001
* Significant difference in comparison to vehicle (* P < 0.05, * * P < 0.01, * * * P < 0.001). All slopes were significant at the level of P < 0.001 Abbreviations as in Table I.
the r e g r e s s i o n lines d e s c r i b i n g b a s e l i n e r e l a t i o n ship b e t w e e n systolic b l o o d p r e s s u r e a n d corres p o n d i n g h e a r t r a t e p e r i o d ( T a b l e II). In e a c h e x p e r i m e n t a l situation t h e slopes w e r e significantly s m a l l e r in S H R t h a n in W K Y . L V injections o f 25, 125 a n d 500 ng of A N P d i d not c h a n g e significantly b l o o d p r e s s u r e or h e a r t r a t e n e i t h e r in W K Y n o r in S H R ( d a t a not shown). T h e highest dose o f A N P r e d u c e d t h e slope of t h e r e g r e s s i o n line d e s c r i b i n g b a s e l i n e relationship between SAP and corresponding H R p in S H R ( T a b l e II).
125 n g
10 n g
ANP
AVP+
125
ng ANP
WKY MAP(mmHg) HR(b/min)
102_+ 3 341+12
104_+ 4 337_+14
105_+ 2 345_+ 7
103+ 3 335+15
SHR MAP(mmHg) HR ( b / m i n )
163_+ 5 390_+23
167_+ 6 373_+14
162_+ 6 395_+24
164_+ 5 398_+ 12
Abbreviations as in Table 1. c o m b i n e d a d m i n i s t r a t i o n o f A N P a n d A V P in W K Y a n d S H R b l o o d p r e s s u r e i n c r e a s e d to 107 + 3 a n d to 170 + 5 m m H g , respectively. T h e s e values w e r e no l o n g e r significant in c o m p a r i s o n to c o n t r o l values. P h e n y l e p h r ine ~
WKY
SHR
loo 80
I E Ck <
4O 2O 0
Interaction of ANP and A VP T a b l e I I I p r e s e n t s m e a n values of M A P a n d H R in W K Y a n d S H R b e f o r e a d m i n i s t r a t i o n o f A C S F , 125 ng A N P , 10 ng A V P a n d 10 ng A V P + 125 ng A N P simultaneously. Similarly as in Series 1, L V a d m i n i s t r a t i o n o f A V P a l o n e res u i t e d in b o t h strains in significant i n c r e a s e s of M A P as c o m p a r e d to c h a n g e s o b s e r v e d after A C S F , while A N P a l o n e was n o t effective. A f t e r injection o f 10 ng of A V P b l o o d p r e s s u r e inc r e a s e d from 104 + 4 to 113 + 4 ( P < 0.01) a n d from 167 + 6 to 178 + 6 ( P < 0.001) m m H g , in W K Y a n d S H R , respectively. In b o t h strains p r e s s o r effects o f A V P w e r e m a r k e d l y a t t e n u a t e d by s i m u l t a n e o u s a d m i n i s t r a t i o n of A N P . A f t e r
10 n g AVP
7o 5O 40 3O 2O <1
10 0 10 -20 acsf
ANP
AVP
AVP+ANP
Fig. 4. Maximum changes of systolic pressure (SAP) (upper panel) and heart rate period (HRp) (lower panel) caused by i.v. injections of phenylephrine in WKY and SHR after LV injection of ACSF, ANP, AVP and combination of AVP with ANP. * Significant difference between WKY and SHR; (* P < 0.05, ** P < 0.01, *** P < 0.001).
39 Sodium n i t r o p r u s s i d e
-2oL
r///~
WKY
SHR
o
-40
-6O O3 --8O
~o.o5
J
-100
--10
decrements in HRp after simultaneous LV administration of ANP and AVP, however, were significantly smaller in SHR than in WKY (Fig. 5). Factorial ANOVA on the slopes coefficients of significant regression lines describing relationships between SAP and HRp during changes of blood pressure induced by Phe and SN revealed significant reduction of the slopes by the following treatments: (1) by AVP in WKY and SHR during SN-induced hypotension and; (2) in SHR by AVP, ANP and combination of ANP and AVP during Phe-induced hypertension (Fig. 6).
-2O
Discussion
-30
"r
<:~
--40 -50 --60
acsf
ANP
AVP
ANP+AVP
Fig. 5. Maximum changes of systolic pressure (SAP) (upper panel) and heart rate period (HRp) (lower panel) caused by i.v. injection of sodium nitroprusside in WKY and SHR after LV injection of ACSF, ANP, AVP and combination of AVP with ANP. For other explanations see Fig. 4
Figs. 4 and 5 present maximum changes of SAP and HRp elicited by systemic administration of phenylephrine and sodium nitroprusside in WKY and SHR after LV administration of ACSF, ANP, AVP and combination of ANP and AVP. Phenylephrine elicited similar maximum increases in SAP and HRp after each treatment in both strains (Fig. 4). In each experimental situation Phe induced significantly smaller increments in HRp in SHR than in WKY. In WKY SN-induced changes of SAP and HRp were not significantly affected by any of the treatments. SN-induced decrements in SAP after LV administration of ANP, AVP and ANP + AVP were significantly greater in SHR than in WKY. In SHR sodium nitroprusside caused greater decrease in systolic pressure when it was injected after combined administration of ANP and AVP than after ACSF alone. Changes in HRp elicited by SN were not significantly affected by any of the treatments neither in WKY nor in SHR. The
The present study indicates that AVP and ANP elicit differential central cardiovascular effects in normotensive and spontaneously hypertensive rats, and supports evidence for their interaction in blood pressure regulation.
Central effects of AVP in WKY and SHR In agreement with other reports on conscious and anesthetized normotensive rats and rabbits [14,19,24] the present results demonstrate clearcut, long lasting pressor responses to LV administration of relatively small doses of AVP in normotensive and spontaneously hypertensive rats. The pressor response to AVP in the latter strain is evoked by the higher dose and was always associated with heart rate acceleration. In both strains blood pressure increase after bolus LV injection of AVP was manifested with a delay of a few minutes (except from 50 ng in SHR which caused immediate increase of MAP) and lasted 30-40 min. As illustrated in Fig. 1 the pressor response to higher doses of AVP in WKY was distinctly biphasic whereas in SHR it was more regular. The reason for the biphasic pattern in WKY is not clear. Central pressor effect of AVP has been attributed to centrally driven enhancement of sympathetic activity [14,26]. Recently Janiak et al. [14] presented evidence that part of the pressor response may be subserved by systemically released vasopressin. It may be hypothesized that the putative systemic release of AVP could
4()
Sodium -
-
~
- -
nitroprusside AMP
- -
-
-
-
-
ANP
AMP
. . . . . .
ANP
36O 32O 280 WKY
240
/ . . . ~.>-" ...~:.- ~ .
SHR
120 80 40 0
.
.
.
.
'
0
.
.
.
.
40
i
.
.
.
80
.
i
.
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.
120
.
,
.
.
.
.
160
,
.
.
.
200
.
,
.
.
.
.
240
,
280
sP (ram H~ Phenylephrine
360
320 280 240
WKY
200 160 120
40 O
"
0
'
'
'
40
.
.
.
.
'
80
.
.
.
.
'
.
120
.
.
.
'
.
.
160
.
.
'
.
200
.
.
.
'
.
240
.
.
.
J
280
SP (rrrn H~
Fig. 6. Regression lines between systolic blood pressure (SAP) and heart rate period (HRp) during increases of blood pressure elicited by phenylephrine (lower panel) and decreases of blood pressure elicited by sodium nitroprusside (upper panel) after LV injections of ACSF, ANP, AVP and combination of AVP and ANP in WKY (four upper lines) and SHR (four lower lines). * Significant difference in slope in comparison to ACSF (* P < 0.05, ** P < 0.01).
account for the delayed elevation of blood pressure and that the time relations between activation of sympathetic system and of AVP releasing neurons may be different in WKY and in SHR.
Alternatively, transient activation of some con]pensatory blood pressure decreasing mechanism, which is deficient in SHR, should be also considered. Several lines of evidence indicate that AVP significantly contributes to baroreflex control of cardiovascular functions [1]. Circulating vasopressin has been reported to facilitate baroreflex, whereas intracranially administered AVP has been shown to suppress baroreflex in conscious, normotensive rats [2,6,11,23,40]. On the other hand LV lysine vasopressin facilitated baroreflex in anesthetized normotensive Wistar rats but not in SHR [13]. In the present study administration of 10 and 50 ng of AVP resulted in significant reductions of slopes of the regression lines describing baseline relationship between SAP and the corresponding HRp in both strains. This effect could result from: (1) enhancement of central drive from cardioexcitatory sympathetic neurones; (2) suppression of the cardiac component of baroreflex; or (3) nonbaroreflex mediated decrease of the vagal tone. The results of Series 3 experiments support evidence for significant interaction of central AVP with the cardiac component of baroreflex. In both strains, LV-administered AVP increased reflex heart rate acceleration during SN-induced hypotension. The differences in HR acceleration caused by sodium nitroprusside were not significant between the both strains, except in case of combined administration of AVP and ANP which elicited significantly smaller HR acceleration in SHR in spite of greater decrease of systolic pressure in this strain. On the other hand, during blood pressure increase elicited by phenylephrine AVP decreased the slope describing the relationship between SAP and HRp in SHR but not in WKY. Several studies indicate significant impairment of baroreflex control of blood pressure in SHR [7,9,22,30]. The present results are in agreement with these observations as shown by significantly smaller changes of HRp at any given level of SAP during blood pressure changes in WKY than in SHR. Differences between WKY and SHR were more accentuated during blood pressure elevations as indicated by results of our Series 3 experiments, showing that rapid Phe-induced SAP increases
41
The present study does not give evidence for important action of central ANP on baseline blood pressure regulation. Previous studies on anesthetized normotensive rats demonstrated significant decreases or increases of blood pressure after local microinjections of ANP to the nucleus tractus solitarius and to the preoptic area, respectively [7,33]. On the other hand, LV injections of ANP in conscious WKY and SHR failed to affect basal cardiovascular parameters [5,12]. ANP per se did not alter sensitivity of the cardiac component of the baroreflex in the hypotensive range of blood pressure changes in WKY and SHR and in hypertensive range of blood pressure in WKY. Our results give however evidence for suppression by centrally applied ANP of the cardiac component of the baroreflex in SHR (see also below).
tion of the sympathetic system and vasopressin release [14]. With this regard, central ANP has been reported to suppress activity of the sympathetic system and AVP release in the rat [15,28,31,37], which could account for reduction of the pressor response to AVP seen in the present study. As shown in our Series 3 experiments, simultaneous administration of ANP and AVP also reversed effects of AVP on the cardiac component of the baroreflex during sodium nitroprusside hypotension in WKY and SHR. Interestingly, in WKY ANP did not significantly modify sensitivity of cardiac component of baroreflex during the induced hypertension while in SHR the sensitivity of the CCB was comparably suppressed by AVP, ANP and simultaneous administration of both peptides. This suggests involvement of a common mechanism in central action of both peptides. In view of generally hypotensive action of ANP its suppressory effect on CCB in SHR is not clear. It may be speculated that the neurochemical lesion underlying hypertension in SHR enables to disclose some central actions of ANP which are normally counterbalanced by other mechanisms. In summary, the present results support evidence that centrally applied vasopressin and atrial natriuretic peptide exert significant differential effects on blood pressure regulation in WKY and SHR. Arginine vasopressin is shown to affect both baseline blood pressure/heart rate relationship and cardiac component of the baroreflex in both strains. On the other hand, atrial natriuretic peptide acts mainly by interaction with AVP in both strains and modulation of the reflex blood pressure control in SHR. Both AVP and ANP appear to impair cardiac component of reflex buffering of blood pressure in SHR.
Interaction of atrial natriuretic peptide with vasopressin
Acknowledgments
In agreement with our earlier data [36] the present study indicates existence of significant central interaction between ANP and AVP in WKY and SHR. In both strains simultaneous administration of AVP and ANP abolished significant increase of blood pressure induced by AVP. Pressor effects of AVP are attributed to activa-
The authors wish to thank Calbiochem-Behring Corp. for the generous gift of arginine-8vasopressin. The generous gift of rat ANP was obtained from Peninsula Inc. thanks to the courtesy of Dr. Andrzej Januszewicz from the Department of Hypertension and Angiology (Medical Academy of Warsaw).
resulted in significantly smaller bradycardia in SHR. The finding that in WKY central AVP did not have any significant effect on reflex bradycardia during Phe-induced hypertension, whereas in SHR it elicited significant suppression of CCB suggests specific impairment of reflex buffering of blood pressure increase in this strain by central arginine vasopressin. The available evidence suggests that circulating and central AVP exerts a double modulatory effect on reflex regulation of blood pressure. Our recent data indicate that circulating V2 agonist potentiate reflex bradycardia in WKY but not in SHR [3]. Taken together these data indicate that in SHR the facilitatory action of AVP on baroreflex is impaired leaving the suppressory effect of central AVP unopposed.
Central effects of atrial natriuretic peptide
42
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Genest. J. and Cantin, M., Effect of synthetic atrial natriuretic factor on arginine vasopressin release by the ral hypothalamo-neurohypophyseal complex in organ culture. Biochem. Biophys. Res. Commun., 134 (1986) 652-658. Lapinski, M., Stepniakowski, K., Januszewicz. A., Noszczyk, B., Szczepanska-Sadowska, E., Atrial ualriuretic factor enhances vasopressin-induced bradycardia in normotensive (WKY) but not in spontaneously hypertensive (SHR) rats, Eur. J. Clin. Invest., 18 (19881 568-574. Manning, P.T., Schwartz, D., Katsube, N.C., Holmberg, S.W., Needleman, P., Vasopressin - stimulated release of atriopeptin: endocrine antagonists in fluid homeostasis, Science, 229 (1985) 395-397. Mantych, Ch.R., Kruger, L., Brecha, N.C. and Mantych, P.W., Localization of specific binding sites for atrial natriuretic factor in the central nervous system of rat, guinea pig, cat and human, Brain Res., 4123 (1987) 329-342. Martin, S.M., Malkinson, T.J., Veale, W.L. and Pittman, Q.J., The action of centrally administered arginine vasopressin on blood pressure in the conscious rabbit, Brain Res. 348 (1985) 137 145. Mohring, J., Schoun, J., Kintz, J., McNeill, R., Decreased vasopressin content in brain stem of rats with spontaneous hypertension, Naunyn-Schmiedeberg's Arch. Pharmacol., 315 (1990) 83-84. Morris, M., Keller, M. and Sundberg, D.K., Changes in paraventricular vasopressin and oxytocin during the development of spontaneous hypertension, Hypertension, 5 (1983) 476-481. Nosaka, S. and Wang, S.C., Carotid sinus baroreceptor functions in the spontaneously hypertensive rat, Am. J. Physiol., 222 (1972) 1079-1089. Peuler, J.D., Edwards, G.L., Schmid, P.G. and Johnson, A.K., Area postrema and differential reflex effects of vasopressin and phenylephrine in rats, Am. J. Physiol. 258 (1990) H1255-H1259. Pittman, Q.J., Lawrence, D. and McLean, L., Central effects of arginine vasopressin on blood pressure in rats, Endocrinology, 110 (1982) 1058-1060. Rascher, W., Lang, R.E., Unger, T., Ganten, D. and Gross, F., Vasopressin in brain of spontaneously hypertensive rats, Am. J. Physiol., 242 (1982) H496-H499. Riphagen, C.L. and Pittman, Q.J., Mechanisms underlying the cardiovascular responses to intrathecal vasopressin administration in rats, Can. J. Physiol. Pharmacx)l., 67 (1989) 269-275. Saavedra, J.M., Israel, A., Kurihara, M. and Fuchs, E., Decreased number and affinity of rat atrial natriuretic peptide (6-33) binding sites in the subfornical organ of spontaneously hypertensive rats, Circ. Res., 58 (1986) 386392. Samson, W.K., Central nervous system actions of atrial natriuretic factor, Brain Res. Bull., 20 (1987) 831-837. Saper, C.B., Standaert, D.G., Currie, M.G., Schwartz, D., Geller, D.M., Needleman, P., Atriopeptin - immunoreacrive neurons in the brain: presence in cardiovascular regulatory areas, Science, 227 (1985) 1047-1049.
43 30 Sapru, H.N., Wang, S.C., Modification of aortic baroreceptor resetting in the spontaneously hypertensive rat, Am. J. Physiol., 230 (1976) 664-670. 31 Schultz, H.D., Steele, M.K., Gardner, D.G., Central administration of atrial peptide decreases sympathetic outflow in rats, Am. J. Physiol., 258 (1990) R1250-R1256. 32 Wiener, B.J., Statistical Principles in Experimental Design. McGraw-Hill, New York, 1970, pp46-139. 33 Sills, M.A., Nguyen, K.Q. and Jacobowitz, D.M., Increases in heart rate and blood pressure produced by microinjections of atrial natriuretic factor into the AV3V region of rat brain, Peptides, 6 (1985) 1037-1042. 34 Sladek, C.D., Devine, M.A., Felten, S.J., Aravich, P.F. and Blair, M.L., Abnormalities in hypothalamic neurohypophysial vasopressin content are not a consequence of hypertension in the spontaneously hypertensive rat, Brain Res., 445 (1988) 39-46. 35 Sladek, J.R., Davis, B.J. and Sladek, C.D., Localization of vasopressin - neurophysin and norepinephrine in the supraoptic nucleus of spontaneously hypertensive rats, Brain Res., 365 (1986) 293-304.
36 Stepniakowski, K., Budzikowski, A., Lon, S. and Szczepanska-Sadowska, E., Central ANP attenuates pressor responses to central AVP in WKY and SHR, Brain Res. Bull., 27 (1991) 247-249. 37 Szczepanska-Sadowska, E., Wocial, B., Sobocinska, J., Kowalik-Borowka, E., Central effects of atrial natriuretic peptide on plasma catecholamines, vasopressin, renin and beta-endorphin and renal excretory functions in the dog, Horm. Metab. Res., 22 (1990) 323-328. 38 Szczepanska-Sadowska, E., Interaction of vasopressin and of the atrial natriuretic peptide in blood pressure control, Acta Physiol. Scand., 136 (1989) (Suppl. 583) 79-87. 39 Tribollet, E., Barberise, C., Jard, S., Dubois-Dauphin, M. and Dreifuss, J.J., Localization and pharmacological characterization of high affinity binding sites for vasopressin and oxytocin in the rat brain by light microscopic autoradiography, Brain Res., 442 (1988) 105-118. 40 Unger, T., Rohmeiss, P., Demmert, G., Ganten, D., Lang, R.E. and Luft, F.C., Differential modulation of the baroreceptor reflex by brain and plasma vasopressin, Hypertension 8 (1986) (Suppl. II): 11157-11162.
33
© 1994 Elsevier Science B.V. All rights reserved 0165-1838/94/$07.00 JANS 01480
Central cardiovascular effects of AVP and ANP in normotensive and spontaneously hypertensive rats Konrad St~pniakowski a, Adam Budzikowski b, S/awomir Loft b and Ewa Szczepafiska-Sadowska b,, a Departments of Hypertension and Angiology and b Department of Clinical and Applied Physiology, MedicalAcademy of Warsaw, 00-927 Warsaw, Poland (Received 14 April 1993) (Revision received 20 June 1993) (Accepted 12 July 1993)
Key words: Vasopressin; Atrial natriuretic peptide; Blood pressure; Central actions; SHR; WKY Abstract The purpose of the present study was to compare influence of central arginine vasopressin (AVP) and of atrial natriuretic peptide (ANP) on control of arterial blood pressure (MAP) and heart rate (HR) in normotensive (WKY) and spontaneously hypertensive (SHR) rats. Three series of experiments were performed on 30 WKY and 30 SHR, chronically instrumented with guide tubes in the lateral ventricle (LV) and arterial and venous catheters. MAP and HR were monitored before and after i.v. injections of either vehicle or 1, 10 and 50 ng of AVP and 25, 125 and 500 ng of ANP. Sensitivity of cardiac component of baroreflex (CCB), expressed as a slope of the regression line was determined from relationships between systolic arterial pressure (SAP) and HR period (HRp) during phenylephrine (Phe)-induced hypertension and sodium nitroprusside (SN)-induced hypotension. CCB was measured before and after administration of either vehicle, AVP, ANP, or both peptides together. Increases of MAP occurred after LV administration of 1, 10 and 50 ng of AVP in WKY and of 10 and 50 ng in SHR. ANP did not cause significant changes in MAP in both strains as compared to vehicle, but it abolished AVP-induced MAP increase in WKY and SHR. CCB was reduced in WKY and SHR after LV administration of AVP during SN-induced hypotension. In SHR but not in WKY administration of ANP, AVP and ANP + AVP decreased CCB during Phe-induced MAP elevation. The results indicate that centrally applied AVP and ANP exert differential effects on blood pressure and baroreflex control of heart rate in WKY and SHR and suggest interaction of these two peptides in blood pressure regulation at the level of central nervous system.
Introduction
Many recent investigations suggest the existence of mutual interactions between atrial natriuretic peptide (ANP) and vasopressin (AVP) in regulation of body fluid balance and blood pressure. Systemically administered ANP has been
* Corresponding author. Department of Clinical and Applied Physiology, Medical Academy of Warsaw, Krakowskie Przedmiescie 26/28 00-927 Warsaw, Poland. Tel.: 26-45-86.
SSDI 0 1 6 5 - 1 8 3 8 ( 9 3 ) E 0 0 9 7 - O
found to reduce blood pressure increase and to enhance reflex bradycardia elicited by systemic vasopressin [16,38]. Immunohistochemical evidence indicates the similar location of ANP and AVP and their receptors in forebrain and brain stem areas suggesting that these peptides contribute to blood pressure regulation [4,10,17,18,28,29,39]. Significant differences in AVP content and density of ANP binding sites in spontaneously hypertensive (SHR) and normotensive (WKY) rats [20,21,25,27,34,35] suggest the role of these two neuropeptides in the
34
pathogenesis of hypertension. The purpose of the present study was to compare blood pressure and heart rate responses to central administration of AVP and ANP as well as the interaction between these two peptides in blood pressure and heart rate regulation in normotensive (WKY) and spontaneously hypertensive (SHR) rats. We have found that central effects of vasopressin and ANP on blood pressure and the sensitivity of cardiac component of baroreflex (CCB) markedly differ in WKY and SHR. Moreover, our results indicate that ANP suppresses AVP-induced pressor responses in WKY and SHR and modifies AVP-induced changes in sensitivity of the cardiac component of baroreflex in WKY.
Material and Methods
Material Experiments were performed on conscious, freely moving SHR and WKY, males, 12-14 weeks old (body weight between 280-300 g), maintained on a 12-h l i g h t / d a r k cycle (light from 07.00 to 19.00 h). Before the experiments they had free access to water and commercial laboratory chow, containing 0.5% NaCI. Food and water were removed during the experimental sessions.
Surgical procedures Seven to 10 days before the experiment the animals were anesthetized with chloral hydrate (chloral-hydrate, Merck, 36 mg/100g i.p.). A stainless steel guide tube (O.D. 0.6 mm) was implanted into the lateral cerebral ventricle (LV) through the opening drilled in the skull according to the following parameters: - 1 . 5 mm posterior to the bregma, 2 mm lateral to the sagittal suture and 5 mm below the surface of the skull. The cannula was closed with a stainless steel stylet and fixed in the skull with acrylic cement. One or 2 days prior to the experiment all rats were subjected to ether anesthesia and instrumented with arterial catheter (PE 20) for blood pressure measurements during freely moving state. The catheter was inserted into the aorta through the left femoral artery. In the third group of animals
the additional PE 20 catheter was introduced into the vena cava through the right femoral vein for injections of phenylephrine (Phe) and sodium nitroprusside (SN). The catheters were connected to PE 60 tubing filled with saline and heparin, plugged with stoppers, tunnelled under the skin and exteriorized on the neck. After the surgery the animals were placed in individual experimental cages for recovery.
Experimental protocol The experiments were performed on 30 WKY and 30 SHR having icv cannulae appropriately located in the LV. The animals were divided into three experimental groups, each consisting of t0 WKY and 10 SHR. Mean values of resting MAP and heart rate (HR) found in the three experimental groups are presented in Tables I and II1. At the beginning of the experiment the arterial catheter was connected to the blood pressure unit for MAP and H R measurements. The stylet was removed from the LV cannula. The actual experiments started after 30-40 min of rest to allow for stabilization of MAP and HR. LV injections were made by means of Hamilton microsyringe at a rate of 5 / z l / 1 0 s. In Series 1 effect of central AVP on MAP and H R was compared in 10 WKY and 10 SHR. At the beginning of each experimental session the animals received LV injection of vehicle (5 pA). This was followed by administration of 1, 10 and 50 ng of AVP in 5 ~1 of vehicle. Subsequent injections were performed with time intervals of at least 40 min when MAP and H R returned to the resting level (Table I). Blood pressure -systolic (SAP), diastolic (DAP) and heart rate were monitored every 1 min during 10 min and every 5 rain during subsequent 30 min following the injection. To assess the effect of central AVP on the relationship between systolic arterial pressure (SAP) and heart rate period (HRp) individual values of SAP and corresponding (beat-to-beat) HRp during the first 10 min after icv administration of either vehicle or AVP were subjected to regression analysis. Slopes of the regression lines significant at the level of P < 0.001 were subjected to A N O V A to estimate significance of differences between treatments and strains.
35 TABLE I Mean blood pressure and heart rate calues preceding L V AVP and ANP injections
Series 1
Vehicle 1 ngAVP
MAP (mmHg) 114_+ 3 113_+ 3 HR (b/ rain) 327_+11 339_+12
10ngAVP
50ngAVP
117_+ 3
117_+ 3
339_+12
351_+ 9
176_+ 3
1795:4
406_+ 1
426_+13
SI-IR
MAP (mmHg) 178_+ 2 175-+ 3 HR (b/ rain) 407-+12 396_+12
Series 2 ACSF 10 ng ANP 125 ng ANP 500 ng ANP WKY MAP (mmHg) 97J: 4 99+ 3 94-+ 3 96+ 3 HR (b/ min) 321+ 8 339+ 9 317_+ 9 334+ 4 SItR
MAP (mmHg) 185_+ 4 185+ 4 HR (b/ min) 402+10 386+12
185_+ 3
188+ 3
4025:11
406+12
Values are as mean+SEM. ACSF, artificial cerebrospinal fluid; AVP, arginine vasopressin, ANP, atrial natriuretic peptide; WKY, normotensive Wistar Kyoto rats; SHR, spontaneously hypertensive rats. Effect of central administration of A N P on M A P and H R was compared in Series 2. The animals received LV injections of either vehicle or with 25, 125 and 500 ng of atrial natriuretic peptide (ANP) in 5/zl of vehicle. The experimental procedure was identical as in Series 1. The relationships between SAP and H R p during the first 10 min after administration of either artificial cerebrospinal fluid (ACSF) or A N P were determined and subjected to regression analysis as in Series 1. Series 3 was conducted to find out whether central A N P and A V P interact in blood pressure regulation. The animals were subjected to LV administrations of (1) vehicle, (2) 125 ng of ANP, (3) 10 ng of AVP and 4) 10 ng AVP + 125 ng ANP, at the minimum of at least 40-min intervals. Ten minutes after each LV injection phenyl-
ephrine (Phe, 0.5 /zg/100 g) was given i.v. as a bolus in 100 /~1 of 0.9% NaCI and its effects on blood pressure and heart rate recorded continuously. When M A P and H R stabilized to the level found prior to Phe administration (usually 5 - 7 min after i.v. Phe injection) the rats were injected i.v. with sodium nitroprusside (SN, 3 /~g/100 g) in 100 /3,1 0.9% NaC1, while blood pressure and heart rate were recorded continuously. After completion of the experiments the position of the cannula in the lateral ventricle was verified by injection of Evans Blue and postm o r t e m brain examination. In some animals the appropriate location of the cannula was preliminary verified by testing presence of drinking and pressor response to LV administration of 100 ng of angiotensin II. Measurements
For blood pressure and heart rate measurements the arterial catheter was connected to the blood pressure recording device consisting of transducer and amplifier (Statham, Gould P 23Db). The signal frequency was modulated with analog to digital converter connected through an interface to Amstrad-Schneider CPC 6128 computer which calculated on line the following parameters: systolic (SAP, mmHg), diastolic (DAP, m m H g ) mean (MAP, m m H g ) pressure and heart rate period (HRp, ms). The latter corresponded to the interval between consecutive SAP. To assess sensitivity of the cardiac component of the baroreflex (CCB) [35] individual regression analysis were performed for relationships between SAP and the following H R p from the onset till the maximum of the pressor and depressor responses to Phe and SN, respectively [8]. Changes in CCB after each treatments were evaluated on the basis of differences in slopes of significant regressions. To this end, individual regression coefficients, significant at a level of no less than P < 0.01 were subjected to factorial A N O V A , with treatments and strains as factors. Drugs
Arginine vasopressin (arginine-8-vasopressin, Calbiochem) was kept frozen as a stock solution in 0.1 M acetic acid and freshly diluted with
3~)
artificial cerebrospinal fluid (ACSF) to the required concentration. Appropriate dilution of 0.1 M acetic acid with ACSF served as vehicle. Atrial natriuretic peptide (rat-28 ANF, Peninsula) was kept lyophilized and freshly diluted with ACSF to the required concentration. The ACSF served as vehicle in this series of experiments. No significant difference was noted in heart rate and pressor responses in WKY and SHR to control injections of the vehicles used in Series 1, 2 and 3. The ACSF contained (mM) Na + - 156.0; K + 3.0; Ca 2 + - 1.5; CI- 147.0; H C O ~ 15.
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Results
Central effects of vasopressm and of atrial natriuretic peptide Mean values of MAP and H R preceding injections of AVP and ANP in WKY and SHR are presented in Table I. Administration of 1, 10 and 50 ng of AVP in WKY and of 10 and 50 ng in SHR elicited significant increases of blood pressure (Figs. 1 and 2). In WKY MAP responses
45
rnin
B
Statistical analysis Means and standard errors are presented in the study. Student's t-test, single factor and factorial analysis of variance (ANOVA) for repeated measurements were applied to appropriate data. In each series consecutive values of MAP and HRp after LV injections were compared to the baseline pre-injection values by single factor A N O V A for repeated measurements. The differences in changes in MAP and H R p between various treatments or between WKY and SHR were evaluated by factorial A N O V A on differences from the preinjection level. The NewmanKeul's a posteriori test or multiple comparisons were used to isolate significant differences between individual means [32]. Changes in MAP and H R p elicited by various treatments at a given time were compared by t-test. Linear analysis of regression was performed by the least squares method. The slopes of the regression lines between groups were compared with ANOVA. The results were considered to be significant if P < 0.05.
~1
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Fig. 1. Mean arterial blood pressure changes after LV injection of vehicle (empty circles) and 1 (A), 10 (B) and 50 (C) ng of A V P (solid circles) in WKY. * Significant difference in A V P vs. vehicle changes; (* P < 0.05, * * P < 0.01, * * * P < 0.001). x Significant difference in comparison to initial value; (x p < 0.05, x× p < 0.01, x x x p < 0.001).
after 10 and 50 ng of AVP were biphasic with peaks at 10 and 30-35 min whereas in SHR the same doses resulted in monophasic prolonged increase of blood pressure. In SHR the lowest dose of AVP elicited small decrease in MAP
37
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in HRp (Fig. 3). Changes of HRp after injection o f 50 ng o f A V P in W K Y and S H R w e r e significantly different from each other by two-way A N O V A I F ( l , 1 8 ) = 7.03; P < 0.05].
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E x x -g xx _1~
x
x
l l l l l l l l l l J l l l l l l l l l l l l l l l l l l l l l l l l t l l l X ~ l l l .0...5
.....
t0 ....
15 . . . .
20 ....
25 ....
30 ....
35 ....
40 ....
45
min <]
-5 -10 •
0
.
.
.
.
5
.
.
.
.
10 .
.
.
.
15 .
.
.
.
20
.
.
.
.
25
.
.
.
.
30
.
.
.
.
35
.
.
.
.
4 0
.
.
.
.
C
45
rain --e-- vehicle
Fig. 2. Mean arterial blood pressure changes after LV injection of vehicle and 1 (A), 10 (B) and 50 (C) ng of AVP in SHR. For other explanations see Fig. 1.
by two-way A N O V A [ F ( 1 , 1 8 ) = 7.17; P < 0.05]. In W K Y H R was not significantly affected,
whereas in SHR injection of 10 and 50 ng of AVP increased HR as shown by significant decreases
ng
AVP
~
which, however, was not different from that observed after LV vehicle in this strain. Changes of MAP after the lowest dose of AVP in WKY and SHR were significantly different from each other
--e-- 5 0
3
- g
-15. o :' i i '51' i iJ i' ','51'' '2'0'." '2'51'. i i;'ii '3'511'i~:ii :,5 rain
Fig. 3. Heart rate period changes after LV injection of 1 (A), 10 (B) and 50 (12) ng of AVP in SHR in comparison to control LV injection of vehicle. For other explanations see Fig. 1.
TABLE II
" F A B L E III
Slopes of the regression lines describing relationship between SAP and HRp under control conditions (Vehich,) and after administration ~# A VP and ANP
Mean blood pressure and heart rate t'alues be~ibre L V inlecttons of A VP and A N P in Series 3 ACSF
Treatment
WKY
SHR
Vehicle AVP I ng 10ng 50ng Vehicle ANP 25 ng 125 ng 5110 ng
1.26
0.71
< 0.001
1.24 1.16 * 1.1 * * * 1.39
0.72 0.65 ** 0.59 * * * 0.67
< 1t.001 <0.001 <0.00t < 0.001
1.35 1.42 1.34
0.66 0.65 0.62 *
P (WKY vs. SHR)
< 0.00l < 0.001 < 0.001
* Significant difference in comparison to vehicle (* P < 0.05, * * P < 0.01, * * * P < 0.001). All slopes were significant at the level of P < 0.001 Abbreviations as in Table I.
the r e g r e s s i o n lines d e s c r i b i n g b a s e l i n e r e l a t i o n ship b e t w e e n systolic b l o o d p r e s s u r e a n d corres p o n d i n g h e a r t r a t e p e r i o d ( T a b l e II). In e a c h e x p e r i m e n t a l situation t h e slopes w e r e significantly s m a l l e r in S H R t h a n in W K Y . L V injections o f 25, 125 a n d 500 ng of A N P d i d not c h a n g e significantly b l o o d p r e s s u r e or h e a r t r a t e n e i t h e r in W K Y n o r in S H R ( d a t a not shown). T h e highest dose o f A N P r e d u c e d t h e slope of t h e r e g r e s s i o n line d e s c r i b i n g b a s e l i n e relationship between SAP and corresponding H R p in S H R ( T a b l e II).
125 n g
10 n g
ANP
AVP+
125
ng ANP
WKY MAP(mmHg) HR(b/min)
102_+ 3 341+12
104_+ 4 337_+14
105_+ 2 345_+ 7
103+ 3 335+15
SHR MAP(mmHg) HR ( b / m i n )
163_+ 5 390_+23
167_+ 6 373_+14
162_+ 6 395_+24
164_+ 5 398_+ 12
Abbreviations as in Table 1. c o m b i n e d a d m i n i s t r a t i o n o f A N P a n d A V P in W K Y a n d S H R b l o o d p r e s s u r e i n c r e a s e d to 107 + 3 a n d to 170 + 5 m m H g , respectively. T h e s e values w e r e no l o n g e r significant in c o m p a r i s o n to c o n t r o l values. P h e n y l e p h r ine ~
WKY
SHR
loo 80
I E Ck <
4O 2O 0
Interaction of ANP and A VP T a b l e I I I p r e s e n t s m e a n values of M A P a n d H R in W K Y a n d S H R b e f o r e a d m i n i s t r a t i o n o f A C S F , 125 ng A N P , 10 ng A V P a n d 10 ng A V P + 125 ng A N P simultaneously. Similarly as in Series 1, L V a d m i n i s t r a t i o n o f A V P a l o n e res u i t e d in b o t h strains in significant i n c r e a s e s of M A P as c o m p a r e d to c h a n g e s o b s e r v e d after A C S F , while A N P a l o n e was n o t effective. A f t e r injection o f 10 ng of A V P b l o o d p r e s s u r e inc r e a s e d from 104 + 4 to 113 + 4 ( P < 0.01) a n d from 167 + 6 to 178 + 6 ( P < 0.001) m m H g , in W K Y a n d S H R , respectively. In b o t h strains p r e s s o r effects o f A V P w e r e m a r k e d l y a t t e n u a t e d by s i m u l t a n e o u s a d m i n i s t r a t i o n of A N P . A f t e r
10 n g AVP
7o 5O 40 3O 2O <1
10 0 10 -20 acsf
ANP
AVP
AVP+ANP
Fig. 4. Maximum changes of systolic pressure (SAP) (upper panel) and heart rate period (HRp) (lower panel) caused by i.v. injections of phenylephrine in WKY and SHR after LV injection of ACSF, ANP, AVP and combination of AVP with ANP. * Significant difference between WKY and SHR; (* P < 0.05, ** P < 0.01, *** P < 0.001).
39 Sodium n i t r o p r u s s i d e
-2oL
r///~
WKY
SHR
o
-40
-6O O3 --8O
~o.o5
J
-100
--10
decrements in HRp after simultaneous LV administration of ANP and AVP, however, were significantly smaller in SHR than in WKY (Fig. 5). Factorial ANOVA on the slopes coefficients of significant regression lines describing relationships between SAP and HRp during changes of blood pressure induced by Phe and SN revealed significant reduction of the slopes by the following treatments: (1) by AVP in WKY and SHR during SN-induced hypotension and; (2) in SHR by AVP, ANP and combination of ANP and AVP during Phe-induced hypertension (Fig. 6).
-2O
Discussion
-30
"r
<:~
--40 -50 --60
acsf
ANP
AVP
ANP+AVP
Fig. 5. Maximum changes of systolic pressure (SAP) (upper panel) and heart rate period (HRp) (lower panel) caused by i.v. injection of sodium nitroprusside in WKY and SHR after LV injection of ACSF, ANP, AVP and combination of AVP with ANP. For other explanations see Fig. 4
Figs. 4 and 5 present maximum changes of SAP and HRp elicited by systemic administration of phenylephrine and sodium nitroprusside in WKY and SHR after LV administration of ACSF, ANP, AVP and combination of ANP and AVP. Phenylephrine elicited similar maximum increases in SAP and HRp after each treatment in both strains (Fig. 4). In each experimental situation Phe induced significantly smaller increments in HRp in SHR than in WKY. In WKY SN-induced changes of SAP and HRp were not significantly affected by any of the treatments. SN-induced decrements in SAP after LV administration of ANP, AVP and ANP + AVP were significantly greater in SHR than in WKY. In SHR sodium nitroprusside caused greater decrease in systolic pressure when it was injected after combined administration of ANP and AVP than after ACSF alone. Changes in HRp elicited by SN were not significantly affected by any of the treatments neither in WKY nor in SHR. The
The present study indicates that AVP and ANP elicit differential central cardiovascular effects in normotensive and spontaneously hypertensive rats, and supports evidence for their interaction in blood pressure regulation.
Central effects of AVP in WKY and SHR In agreement with other reports on conscious and anesthetized normotensive rats and rabbits [14,19,24] the present results demonstrate clearcut, long lasting pressor responses to LV administration of relatively small doses of AVP in normotensive and spontaneously hypertensive rats. The pressor response to AVP in the latter strain is evoked by the higher dose and was always associated with heart rate acceleration. In both strains blood pressure increase after bolus LV injection of AVP was manifested with a delay of a few minutes (except from 50 ng in SHR which caused immediate increase of MAP) and lasted 30-40 min. As illustrated in Fig. 1 the pressor response to higher doses of AVP in WKY was distinctly biphasic whereas in SHR it was more regular. The reason for the biphasic pattern in WKY is not clear. Central pressor effect of AVP has been attributed to centrally driven enhancement of sympathetic activity [14,26]. Recently Janiak et al. [14] presented evidence that part of the pressor response may be subserved by systemically released vasopressin. It may be hypothesized that the putative systemic release of AVP could
4()
Sodium -
-
~
- -
nitroprusside AMP
- -
-
-
-
-
ANP
AMP
. . . . . .
ANP
36O 32O 280 WKY
240
/ . . . ~.>-" ...~:.- ~ .
SHR
120 80 40 0
.
.
.
.
'
0
.
.
.
.
40
i
.
.
.
80
.
i
.
.
.
120
.
,
.
.
.
.
160
,
.
.
.
200
.
,
.
.
.
.
240
,
280
sP (ram H~ Phenylephrine
360
320 280 240
WKY
200 160 120
40 O
"
0
'
'
'
40
.
.
.
.
'
80
.
.
.
.
'
.
120
.
.
.
'
.
.
160
.
.
'
.
200
.
.
.
'
.
240
.
.
.
J
280
SP (rrrn H~
Fig. 6. Regression lines between systolic blood pressure (SAP) and heart rate period (HRp) during increases of blood pressure elicited by phenylephrine (lower panel) and decreases of blood pressure elicited by sodium nitroprusside (upper panel) after LV injections of ACSF, ANP, AVP and combination of AVP and ANP in WKY (four upper lines) and SHR (four lower lines). * Significant difference in slope in comparison to ACSF (* P < 0.05, ** P < 0.01).
account for the delayed elevation of blood pressure and that the time relations between activation of sympathetic system and of AVP releasing neurons may be different in WKY and in SHR.
Alternatively, transient activation of some con]pensatory blood pressure decreasing mechanism, which is deficient in SHR, should be also considered. Several lines of evidence indicate that AVP significantly contributes to baroreflex control of cardiovascular functions [1]. Circulating vasopressin has been reported to facilitate baroreflex, whereas intracranially administered AVP has been shown to suppress baroreflex in conscious, normotensive rats [2,6,11,23,40]. On the other hand LV lysine vasopressin facilitated baroreflex in anesthetized normotensive Wistar rats but not in SHR [13]. In the present study administration of 10 and 50 ng of AVP resulted in significant reductions of slopes of the regression lines describing baseline relationship between SAP and the corresponding HRp in both strains. This effect could result from: (1) enhancement of central drive from cardioexcitatory sympathetic neurones; (2) suppression of the cardiac component of baroreflex; or (3) nonbaroreflex mediated decrease of the vagal tone. The results of Series 3 experiments support evidence for significant interaction of central AVP with the cardiac component of baroreflex. In both strains, LV-administered AVP increased reflex heart rate acceleration during SN-induced hypotension. The differences in HR acceleration caused by sodium nitroprusside were not significant between the both strains, except in case of combined administration of AVP and ANP which elicited significantly smaller HR acceleration in SHR in spite of greater decrease of systolic pressure in this strain. On the other hand, during blood pressure increase elicited by phenylephrine AVP decreased the slope describing the relationship between SAP and HRp in SHR but not in WKY. Several studies indicate significant impairment of baroreflex control of blood pressure in SHR [7,9,22,30]. The present results are in agreement with these observations as shown by significantly smaller changes of HRp at any given level of SAP during blood pressure changes in WKY than in SHR. Differences between WKY and SHR were more accentuated during blood pressure elevations as indicated by results of our Series 3 experiments, showing that rapid Phe-induced SAP increases
41
The present study does not give evidence for important action of central ANP on baseline blood pressure regulation. Previous studies on anesthetized normotensive rats demonstrated significant decreases or increases of blood pressure after local microinjections of ANP to the nucleus tractus solitarius and to the preoptic area, respectively [7,33]. On the other hand, LV injections of ANP in conscious WKY and SHR failed to affect basal cardiovascular parameters [5,12]. ANP per se did not alter sensitivity of the cardiac component of the baroreflex in the hypotensive range of blood pressure changes in WKY and SHR and in hypertensive range of blood pressure in WKY. Our results give however evidence for suppression by centrally applied ANP of the cardiac component of the baroreflex in SHR (see also below).
tion of the sympathetic system and vasopressin release [14]. With this regard, central ANP has been reported to suppress activity of the sympathetic system and AVP release in the rat [15,28,31,37], which could account for reduction of the pressor response to AVP seen in the present study. As shown in our Series 3 experiments, simultaneous administration of ANP and AVP also reversed effects of AVP on the cardiac component of the baroreflex during sodium nitroprusside hypotension in WKY and SHR. Interestingly, in WKY ANP did not significantly modify sensitivity of cardiac component of baroreflex during the induced hypertension while in SHR the sensitivity of the CCB was comparably suppressed by AVP, ANP and simultaneous administration of both peptides. This suggests involvement of a common mechanism in central action of both peptides. In view of generally hypotensive action of ANP its suppressory effect on CCB in SHR is not clear. It may be speculated that the neurochemical lesion underlying hypertension in SHR enables to disclose some central actions of ANP which are normally counterbalanced by other mechanisms. In summary, the present results support evidence that centrally applied vasopressin and atrial natriuretic peptide exert significant differential effects on blood pressure regulation in WKY and SHR. Arginine vasopressin is shown to affect both baseline blood pressure/heart rate relationship and cardiac component of the baroreflex in both strains. On the other hand, atrial natriuretic peptide acts mainly by interaction with AVP in both strains and modulation of the reflex blood pressure control in SHR. Both AVP and ANP appear to impair cardiac component of reflex buffering of blood pressure in SHR.
Interaction of atrial natriuretic peptide with vasopressin
Acknowledgments
In agreement with our earlier data [36] the present study indicates existence of significant central interaction between ANP and AVP in WKY and SHR. In both strains simultaneous administration of AVP and ANP abolished significant increase of blood pressure induced by AVP. Pressor effects of AVP are attributed to activa-
The authors wish to thank Calbiochem-Behring Corp. for the generous gift of arginine-8vasopressin. The generous gift of rat ANP was obtained from Peninsula Inc. thanks to the courtesy of Dr. Andrzej Januszewicz from the Department of Hypertension and Angiology (Medical Academy of Warsaw).
resulted in significantly smaller bradycardia in SHR. The finding that in WKY central AVP did not have any significant effect on reflex bradycardia during Phe-induced hypertension, whereas in SHR it elicited significant suppression of CCB suggests specific impairment of reflex buffering of blood pressure increase in this strain by central arginine vasopressin. The available evidence suggests that circulating and central AVP exerts a double modulatory effect on reflex regulation of blood pressure. Our recent data indicate that circulating V2 agonist potentiate reflex bradycardia in WKY but not in SHR [3]. Taken together these data indicate that in SHR the facilitatory action of AVP on baroreflex is impaired leaving the suppressory effect of central AVP unopposed.
Central effects of atrial natriuretic peptide
42
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