Extract
Note: Please read the complete
full text with Figures and Tables at
Introduction
Arterial baroreflex (ABR) is one of the most important regulatory mechanisms in the cardiovascular system. Baroreflex
function, expressed as baroreflex sensitivity (BRS), is an important determinant for many cardiovascular diseases. Clinically,
it has been proven that ABR dysfunction is associated with sudden death in patients with acute myocardial infarction and
mortality in patients with congestive heart
failure[1-4]. Our previous works demonstrated that ABR function was closely
related to atheroscleosis and end-organ damage in
hypertension[5,6].
Furthermore, ABR function also plays an important role
in the onset and prognosis of stroke. There is established
evidence of abnormal ABR function in both animal
models of stroke[7,8] and patients with acute
stroke[9-11]. Recently, a study within our department indicated that ABR function affected
the survival time of stroke-prone spontaneously-hypertensive rats (SHR-SP). The study also found that restoring impaired
ABR function could prevent stroke in hypertension, suggesting that BRS is a new and important predictor for stroke
incidence and that the restoration of ABR function is a new target for the prevention of
stroke[12]. Since ABR function is associated closely with stroke, it is of great clinical
importance to discover drugs with the ability to improve impaired
ABR function.
Clonidine and moxonidine, representing the first and
second generation of antihypertensive drugs, act on the central
nervous system. It was demonstrated that clonidine
improved ABR function in conscious mice through
parasympathetic activation[13]. However, little is known about the
effect of moxonidine. Both folic acid (vitamin
B9) and mecobalamin (methyl-vitamin
B12) belong to the vitamin B group. During the past decade, interest in the health
benefits of these has increased considerably. Recent interest of
B vitamin focused on their beneficial effects on preventing
cardiovascular diseases. Clinical studies indicated that
folic acid improved ABR function in hypertensive
patients[14]. In many cases, folic acid involved biochemical processes that
required the participation of vitamin
B12[15]. Therefore, we
postulated that mecobalamin might improve ABR function
in conscious SHR-SP.
This study was designed to examine the effects of
clonidine, moxonidine, folic acid, and mecobalamin on
impaired ABR function in SHR-SP and the possible
mechanisms involved.
Materials and methods
Animals and drugs Female SHR-SP rats, aged
20-24 weeks, were provided by the Animal Center of the Second
Military Medical University (Shanghai, China). The rats
were housed under controlled temperatures (23_25 °C) and
lighting (8:00_20:00 light, 20:00_8:00 dark) with free
access to food and tap water. All the animals used in the
experiment received humane care in compliance with
institutional guidelines for health and care of experimental
animals.
Clonidine (Sigma Chemical Co, St Louis, MO, USA),
moxonidine (Yahu Pharmaceutical Co, Shanghai, China), folic
acid (Fushu Pharmaceutical Co, Shanghai, China), mecobalamin (Shenyang Wanlong Pharmaceutical Co,
Shenyang, China). All were dissolved with sterile saline and
stored at 0_4 °C.
Blood pressure and heart period
measurement Systolic blood pressure (SBP) and diastolic blood pressure (DBP),
and heart period (HP) were continuously recorded using a
previously described technique[16,17]. Briefly, the rats were
anesthetized with a combination of ketamine (50 mg/kg, ip)
and diazepam (5 mg/kg, ip). A floating polyethylene catheter
was inserted into the lower abdominal aorta via the left
femoral artery for blood pressure (BP) measurement. Another
catheter was placed into the left femoral vein for BRS
measurement. A stomach catheter was inserted when
intragastric (ig) administration was needed or a stainless steel
cannula (0.7 mm OD) was stereotaxically (1.0 mm posterior to
bregma, 1.6 mm lateral from the midline, 4.0 mm below the
surface of the skull) implanted into the lateral cerebral
ventricle for intracerebroventricular (icv) injection. All the
catheters were exteriorized through the interscapular skin. After
a 2 d recovery period, the animals were placed in individual
cylindrical cages containing food and water for BP recording.
The aortic catheter was connected to a BP transducer via a
rotating swivel that allowed the animals to move freely in the
cage. After about 4 h habituation, the BP signal was
digitized by a microcomputer and beat-to-beat SBP, DBP, and
HP values were determined online. The mean values during
a period of 30 min were calculated and served as the SBP,
DBP, and HP.
Baroreflex sensitivity measurement BRS was
determined using a previously described
method[18]. The principle of this method was to measure the prolongation of
HP in response to an elevation of BP. Briefly, a bolus
injection of phenylephrine (2_5 mg/kg) was used to induce
an elevation of SBP between 20_40 mmHg. HP was
plotted against SBP for a linear regression analysis and the slope
of SBP_HP was expressed as BRS (ms/mmHg). The mean
value of 2 measurements with the proper dose served as
the final result.
Experimental protocol Eighty one SHR-SP were
randomly divided into 7 groups. Four groups were designated
for the ig administration of clonidine (1.0 and 10.0 µg/kg),
moxonidine (0.1 and 1.0 mg/kg), folic acid (1.0 mg/kg),
and mecobalamin (1.0 mg/kg). Three groups were used for
the icv injection of clonidine (4 µg/4 µL), moxonidine
(5 µg/4 µL), and mecobalamin (20 µg/4 µL). After 4 h
habitua-tion, basal (predrug) BP was recorded continuously for 30
min and BRS was determined. Thereafter, a single dose of
drug was given via an ig catheter or stainless steel cannula
as designated. One hour (ig moxonidine) or 30 min (the other
drugs) later, the BP was recorded for another 30 min and BRS
was determined again. The mean values of SBP, DBP, HP
and BRS served as post-drug values. For the ig
administration of clonidine or moxonidine, 2 doses were conducted in 1
rat, that is, the lower dose was administrated and post-drug
values were calculated; 30 min later, the higher dose was
administered and the same protocol was performed.
Statistical analysis All data are expressed as mean±SEM.
Statistical analysis was performed with Student's paired
t-test. P<0.05 was considered statistically significant.
Results
Effects of clonidine on BP, HP, and BRS in SHR-SP
Figure 1 illustrates the effects of clonidine (ig) on BP, HP,
and BRS in SHR-SP. SBP and DBP were significantly
decreased (_16±3 mmHg and _12±3 mmHg) and HP was
prolonged dramatically by 10.0 µg/kg clonidine. DBP was
significantly decreased after 1.0 µg/kg clonidine while SBP
and HP were not obviously affected. BRS was enhanced
markedly from 0.30±0.03 ms/mmHg to 0.43±0.04
ms/mmHg by 1.0 µg/kg clonidine and from
0.30±0.03 ms/mmHg to 0.53±0.09 ms/mmHg by 10.0 µg/kg clonidine.
Clonidine (4 µg, icv; Figure 2) significantly decreased
SBP and DBP (_21±3 mmHg and _15±4 mmHg), but HP was
not significantly prolonged. BRS markedly increased
from 0.30±0.02 ms/mmHg to 0.55±0.09 ms/mmHg.
Effects of moxonidine on BP, HP and BRS in SHR-SP
As shown in Figure 3, both SBP and DBP were significantly
decreased by 2 doses of moxonidine (ig). The reduction in
SBP was 10±2 mmHg by 0.1 mg/kg and 17±3 mmHg by 1.0
mg/kg moxonidine. HP was significantly prolonged by 1.0
mg/kg moxonidine. BRS was significantly improved
(0.27±0.03 vs 0.43±0.03 ms/mmHg and 0.27±0.03
vs 0.54±0.09 ms/mmHg) by 0.1 and 1.0 mg/kg moxonidine (ig).
The effects of moxonidine (icv) are shown in Figure 4.
Moxonidine (5 µg) significantly decreased SBP (189±7
vs 166±5 mmHg) and DBP (132±7
vs 108±4 mmHg). HP was markedly prolonged. BRS was significantly enhanced from
0.38±0.04 ms/mmHg to 0.68±0.05 mmHg. These results
indicated that clonidine and moxonidine improved ABR
function through central mechanisms.
Effects of mecobalamin on BP, HP, and BRS in SHR-SP
Figure 5 shows that 1.0 mg/kg mecobalamin (ig) did not cause
changes in SBP, DBP, and HP. BRS was increased
significantly (0.33±0.02 vs 0.44±0.04 ms/mmHg). Similar to the ig
administration, the icv administration of 20 µg mecobalamin
(Figure 6) did not affect SBP, DBP, and HP. However, BRS
was not changed by icv injection. These results revealed
that mecobalamin increased BRS through peripheral mechanisms.
Effects of folic acid on BP, HP, and BRS in SHR-SP
Similar to the ig administration of mecobalamin, 1.0
mg/kg of ig-administered folic acid (Figure 7) increased BRS
without influence on the SBP, DBP, and HP levels. Icv
injection was not conducted because folic acid could not reach
the concentration needed in this study.
Discussion
The present study clearly demonstrated that clonidine,
moxonidine, folic acid, and mecobalamin all improved the
damaged ABR function in SHR-SP, whether or not the BP
levels were changed or unchanged. It is well known that
ABR function, damaged in many cardiovascular diseases,
including myocardial infarction, heart failure,
athero-sclerosis, diabetes, and end-organ damage in hypertension,
plays a key role in the regulation of cardiovascular acti-vities.
Since the end of 1980s, the pathological importance of ABR
function has attracted the attention of many investigators.
Nowadays, more and more studies have demonstrated that
ABR function is closely associated with stroke. Robinson
et al reported a significant reduction in cardiac BRS
after acute stroke in
patients[10]. Thereafter, they reported that
post-stroke patients with impaired BRS values
(£5.0 ms/mmHg) had a significantly poorer prognosis than patients
without impaired BRS (>5.0 ms/mmHg). Based on these
results, they suggested therapeutic strategies for stroke by
increasing BRS activity with drugs[11].
Given the clinical importance of BRS in stroke therapy,
we believe it is necessary to find drugs that are effective in
restoring damaged ABR function in stroke patients. The
application of these drugs for the prevention of stroke
incidence may be a new strategy in stroke therapy. In this study,
SHR-SP were used. It was reported that SHR-SP had
damaged ABR function compared with normotensive
Wistar_Kyoto rats[19]. At the same time, SHR-SP are a useful
experimental model for examining the pathogenesis of stroke as
well as their treatment because the cerebrovascular lesions
in these animals are similar to that in
humans[20].
Clonidine, a centrally_acting, antihypertensive drug, is
an agonist for both I1-imidazoline receptors and
α2-adreno-ceptors. Tank et al reported that clonidine improved
spontaneous BRS in conscious mice[13]. In this study, we found
that clonidine enhanced BRS significantly after ig
administration in SHR-SP. To determine the possible mechanism
involved in this effect, icv-administered clonidine was
conducted. The results indicated that icv-administered
clonidine also improved BRS significantly in SHR-SP. These
results suggested that the BRS-improving effect induced by
clonidine was mediated through the central mechanism.
Furthermore, 1.0 µg/kg clonidine did not decrease SBP, but
increased BRS significantly, indicating that this
BRS-improving effect was not secondary to a decrease in BP.
Moxonidine, an agent of the second generation,
antihypertensive drugs acting on the central nervous system,
mediates hypotensive effects by activating central
I1-imidazoline receptors and subsequently decreasing sympathetic nerve
activity[21]. In the present study, the effect of moxonidine on
ABR function in conscious SHR-SP was tested. Our study
was the first to find that ig_administered moxonidine
significantly increased BRS in SHR-SP. Furthermore,
icv-administered moxonidine was performed and the results indicated
that BRS was also enhanced significantly, demonstrating
that the central mechanism was involved in the
BRS-improving effect of moxonidine. However, both 0.1 and 1.0 mg/kg
moxonidine decreased SBP and increased BRS significantly,
so it is unclear whether this effect was dependent on the
decrease of blood pressure or not. Further studies are needed
to testify this mechanism.
Folic acid was considered to have potential protection
against cardiovascular diseases due to its
homocysteine-lowering effect[22]. It was also suggested that folate might
have a direct antioxidant role in vivo, which was
independent of any indirect effects through the lowering of
homocysteine levels[23]. Abundant attention has been focused on
folic acid with its role in the prevention of cardiovascular
disease[24,25]. Bechir et
al[14] proved that folic acid improved
impaired BRS in hypertensive patients. They speculated
that the positive effects of folic acid seemed to be mediated
by a reduction of oxidative stress because oxidative stress
directly interfered with nerve endings of baroneurons in the
arterial wall to damage ABR
function[26]. They also found that folic acid had antioxidative
properties[27]. In the present study, we examined effect of folic acid on ABR function in
SHR-SP. It was found that BRS was significantly enhanced
after ig administration of folic acid under the condition that
both BP and HP were unchanged. The BRS-improving
effect of folic acid in SHR-SP might be conducted through
its antioxidative properties. As well as this, folic acid was
reported to enhance endothelial function and increase the
production of nitric oxide (NO)[28,29]; NO played an
important role in the regulation of ABR
function[30]. This might be another contribution to BRS improvement induced by
folic acid. In this study, an icv injection was not conducted
because folic acid could not reach the concentration needed
in this study. Therefore, it was unclear whether the central
mechanism was involved in this BRS-improving effect and
further studies are needed.
Mecobalamin is one of the active analogs of vitamin
B12. It is the essential cofactor for methionine synthase.
Deficiency in folic acid and vitamin
B12 leads to the elevation of the plasma homocysteine level, which is considered an
independent risk factor in the pathogenesis of
atherosclerosis[31], acute myocardial
infarction[32], stroke[33], and
hypertension[34]. All of these diseases were characterized by poor ABR
function. We speculated that vitamin B12
might affect BRS. In the present study, the effect of vitamin
B12 on ABR function was examined in SHR-SP. It was found that
ig-administered mecobalamin increased BRS markedly, but
icv-administered mecobalamin did not. These results suggested that
the central mechanism was not involved in this
BRS-improving effect. The exact mechanism of this effect by vitamin
B12 is still unclear. Visontai et al
reported a negative correlation between the homocysteine concentration and BRS in
exfoliation syndrome or exfoliation glaucoma
patients[35]. At the same time, folic acid and vitamin
B12 decreased blood plasma
homocysteine[36]. So we postulated that the reduction of the
homocysteine concentration might contribute to the
BRS-improving effect of folic acid and mecobalamin.
In conclusion, this study is the first to directly
demonstrate that clonidine, moxonidine, folic acid, and
meco-balamin all improve impaired BRS in SHR-SP. The central
mechanism was involved in this effect of either clonidine
or moxonidine and mecobalamin improved ABR function
through the peripheral mechanism.
References
1 La Rovere MT, Specchia G, Mortara A, Schwartz PJ. Baroreflex
sensitivity, clinical correlates and cardiovascular mortality among
patients with a first myocardial infarction: a prospective study.
Circulation 1988; 78: 816_24.
2 La Rovere MT, Bigger JT Jr, Marcus FI, Mortara A, Schwartz PJ.
Baroreflex sensitivity and heart-rate variability in prediction of
total cardiac mortality after myocardial infarction. Lancet 1998;
351: 478_84.
3 Mortara A, La Rovere MT, Pinna GD, Prpa A, Maestri R, Febo
Q, et al. Arterial baroreflex modulation of heart rate in chronic
heart failure: clinical and hemodynamic correlates and
prognostic implications. Circulation 1997; 96: 3450_8.
4 Fukuma Y, Munakata K, Fukuma N, Kishida H, Hayakawa H,
Takano T. Correlation between atrial natriuretic peptide and
baroreflex sensitivity in patients with congestive heart failure.
Jpn Circ J 1999; 63: 893_9.
5 Shan ZZ, Dai SM, Su DF. Relationship between baroreceptor
reflex function and end-organ damage in spontaneously
hypertensive rats. Am J Physiol 1999; 277: H1200_6.
6 Cai GJ, Miao CY, Xie HH, Lu LH, Su DF. Arterial baroreflex
dysfunction promotes atherosclerosis in rats. Atherosclerosis
2005; 183: 41_7.
7 Doba N, Reis D. Role of the cerebellum and the vestibular
apparatus in regulation of orthostatic reflexes in the cat. Circ Res
1974; 34: 9_18.
8 Cechetto D, Wilson J, Smith K, Wolski D, Silver MD, Hachinski
VC. Autonomic and myocardial changes in middle cerebral
artery occlusion: stroke models in the rat. Brain Res 1989; 502:
296_305.
9 Eames PJ, Blake MJ, Dawson SL, Panerai RB, Potter JF.
Dynamic cerebral autoregulation and beat to beat blood pressure
control are impaired in acute ischaemic stroke. J Neurol Neurosury
Psychiatry 2002; 72: 467_72.
10 Robinson TG, James M, Youde J, Panerai R, Potter J. Cardiac
baroreceptor sensitivity is impaired after acutestroke. Stroke
1997; 28: 1671_6.
11 Robinson TG, Dawson SL, Eames PJ, Panerai RB, Potter JF.
Cardiac baroreceptor sensitivity predicts long-term outcome
after acute ischaemic stroke. Stroke 2003; 34: 705_12.
12 Liu AJ, Ma XJ, Shen FM, Liu JG, Chen H, Su DF. Arterial
baroreflex: a novel target for preventing stroke in rat
hyperten-sion. Stroke 2007; 38: 1916-23.
13 Tank J, Jordan J, Diedrich A, Obst M, Plehm R, Luft FC,
et al. Clonidine improves spontaneous baroreflex sensitivity in
conscious mice through parasympathetic activation. Hypertension
2004; 43: 1042_7.
14 Bechir M, Enseleit F, Chenevard R, Muntwyler J, Luscher TF,
Noll G. Folic acid improves baroreceptor sensitivity in
hyperten-sion. J Cardiovasc Pharmacol 2005; 45: 44_8.
15 Mangoni AA, Jackson SH. Homocysteine and cardiovascular
disease: current evidence and future prospects. Am J Med 2002;
112: 556_65.
16 Su DF, Cerutti C, Barres C, Vincent M, Sassard J. Blood pressure
and baroreflex sensitivity in conscious hypertensive rats of Lyon
strain. Am J Physiol 1986; 251: H1111_7.
17 Miao CY, Xie HH, Yu H, Chu ZX, Su DF. Ketanserin stabilizes
blood pressure in conscious spontaneously hypertensive rats. Clin
Exp Pharmacol Physiol 2003; 30: 189_93.
18 Su DF, Chen L, Kong XB, Cheng Y. Determination of arterial
baroreflex blood pressure control in conscious rats. Acta
Pharmacol Sin 2002; 23: 103_9.
19 Head GA, Minami N. Importance of cardiac, but not vascular,
hypertrophy in the cardiac baroreflex deficit in spontaneously
hypertensive and stroke-prone rats. Am J Med 1992; 92: S54_9.
20 Yamori Y, Okamoto K. Spontaneous hypertension in the rat: A
model for human "essential" hypertension. Verh Dtsch Ges Inn
Med 1974; 80: 168_70.
21 Armah BI, Hofferber E, Stenzel W. General pharmacology of
the novel centrally acting antihypertensive agent moxonidine.
Arzneimittel-Forsch 1988; 38: 1426_34.
22 Nygard O, Nordrehaug JE, Refsum H, Ueland PM, Farstad M,
Vollset SE. Plasma homocysteine levels and mortality in
patients with coronary artery disease. New Engl J Med 1997; 337:
230_6.
23 Nakano E, Higgins JA, Powers HJ. Folate protects against
oxidative modification of human LDL. Br J Nutr 2001; 86: 637_9.
24 Hernandez-Diaz S, Werler MM, Louik C, Mitchell AA. Risk of
gestational hypertension in relation to folic acid
supplementation during pregnancy. Am J Epidemiol 2002; 156: 806_12.
25 Graham IM, O'Callaghan P. The role of folic acid in the
prevention of cardiovascular disease. Curr Opin Lipidol 2000; 6:
577_87.
26 Chapleau MW, Cunningham JT, Sullivan MJ, Wachtel RE,
Abbound FM. Structural versus functional modulation of the
arterial baroreflex. Hypertension 1995; 26: 341_7.
27 Verhaar MC, Wever RM, Kastelein JJ, van Loon D, Milstien S,
Koomans HA, et al. Effects of oral folic acid supplementation
on endothelial function in familial hypercholesterolemia. A
randomized placebo-controlled trial. Circulation 1999; 100:
335_8.
28 Chambers JC, Ueland PM, Obeid OA, Wrigley J, Refsum H,
Kooner JS. Improved vascular endothelial function after oral B
vitamins: An effect mediated through reduced concentrations of
free plasma homocysteine. Circulation 2000; 102: 2479_83.
29 Doshi SN, McDowell IF, Moat SJ, Payne N, Durrant HJ, Lewis
MJ, et al. Folate improves endothelial function in coronary
artery disease: an effect mediated by reduction of intracellular
superoxide? Arterioscler Thromb Vasc Biol 2001; 21: 1196_202.
30 Nightingale AK, Blackman DJ, Field R, Glover NJ, Pegge N,
Mumford C, et al. Role of nitric oxide and oxidative stress in
baroreceptor dysfunction in patients with chronic heart failure.
Clin Sci 2003; 104: 529_35.
31 Kerkeni M, Addad F, Chauffert M, Chuniaud L, Miled A, Trivin
F, et al. Hyperhomocysteinemia, paraoxonase activity and risk
of coronary artery disease. Clin Biochem 2006; 39: 821_5.
32 Banu S, Mollah FH, Alam MK, Rahman MA, Hamid MA, Wahab
MA, et al. Serum homocysteine concentration in patients with
acute MI and chronic IHD. Mymensingh Med J 2005; 14: 54_7.
33 Torbus-Lisiecka B, Bukowska H, Jastrzebska M, Chelstowski K,
Honczarenko K, Naruszewicz M. Lp(a), homocysteine and a
family history of early ischemic cerebral stroke. Nutr Metab
Cardiovasc Dis 2001; 11: 52_9.
34 Kahleova R, Palyzova D, Zvara K, Zvarova J, Hrach K,
Novakova I, et al. Essential hypertension in adolescents: Association with
insulin resistance and with metabolism of homocysteine and
vitamins. Am J Hypertens 2002; 15: 857_64.
35 Visontai Z, Merisch B, Kollai M, Hollo G. Increase of carotid
artery stiffness and decrease of baroreflex sensitivity in
exfoliation syndrome and glaucoma. Br J Ophthalmol 2006; 90: 563_7.
36 Tungkasereerak P, Ong-ajyooth L, Chaiyasoot W, Ong-ajyooth
S, Leowattana W, Vasuvattakul S, et
al. Effect of short-term folate and vitamin B supplementation on blood homocysteine
level and carotid artery wall thickness in chronic hemodialysis
patients. J Med Assoc Thai 2006; 89: 1187_93.
|
