EJCTS Click here for details of sales representative
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to Personal Folders
Right arrow Download to citation manager
Right arrow Permission Requests
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Novella, S.
Right arrow Articles by Benedito, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Novella, S.
Right arrow Articles by Benedito, S.
Related Collections
Right arrow Cardiac - pharmacology
Right arrow Coronary disease
Right arrow Extracorporeal circulation
Right arrow Great vessels

Eur J Cardiothorac Surg 2007;32:69-76. doi:10.1016/j.ejcts.2007.03.047
Copyright © 2007, European Association for Cardio-Thoracic Surgery. Published by Elsevier B.V. All rights reserved

Plasma levels and vascular effects of vasopressin in patients undergoing coronary artery bypass grafting

Susana Novellaa, Ana Cristina Martíneza, Rosa María Pagána, Medardo Hernándeza, Albino García-Sacristána, Angel González-Pintob, José María González-Santosb, Sara Beneditoa,*

a Departamento de Fisiología, Facultad de Farmacia, Universidad Complutense, 28040 Madrid, Spain
b Departamento de Cirugía Cardiovascular, Hospital General Universitario Gregorio Marañón, 28007 Madrid, Spain

Received 15 January 2007; received in revised form 29 March 2007; accepted 30 March 2007.

* Corresponding author. Address: Sección Departamental de Fisiología, Facultad de Farmacia, Universidad Complutense, 28040 Madrid, Spain. Tel.: +34 91 394 1696; fax: +34 91 394 1696. (Email: sbenedi{at}farm.ucm.es).


   Abstract
Top
 Abstract
1. Introduction
 2. Patients and methods
 3. Results
 4. Discussion
 References
 
Objective: Recent studies have suggested that endogenous vasopressin (AVP) acts as a spasmogen during coronary artery bypass grafting (CABG). Given that AVP could induce vasospasm in the grafted vessel, we assessed the release of this peptide during and after CABG, and explored ways of counteracting its contractile effect on the internal mammary artery (IMA). Methods: Plasma levels of AVP were determined by radioimmunoassay in 16 patients before, during and after CABG. Using isometric force recording techniques, we also investigated the mechanisms involved in the contractile effect of AVP in ring preparations of IMA specimens taken from 95 patients. Results: Plasma AVP levels peaked after the start of cardiopulmonary bypass (CPB) and correlated well with serum osmolality (Pearson's r = 0.9490; P < 0.0001; n = 16). An inverse correlation was observed between plasma AVP levels recorded at this stage and the maximal contraction induced in vitro by AVP in vascular rings from the same patients (Pearson's r = –0.6968; P < 0.01; n = 16). No change in the AVP response was produced by endothelium removal, exposure to the NO precursor (3 x 10–4 M L-arginine), inhibition of nitric oxide (NO) synthase (3 x 10–5 M L-NAME) or soluble guanylate cyclase (3 x 10–6 M 1H-[1,2,4]oxadiazol [4,3,-{alpha}]quinoxalin-1-one (ODQ)), removal of the superoxide anion (100 U/ml superoxide dismutase (SOD) plus 1200 U/ml catalase) or hydroxyl radical (10–4 M deferoxamine), or specific {alpha}1- (10–6 M prazosin) or endothelin (10–5 M bosentan) receptor antagonism. In contrast, adenylate cyclase activation (3 x 10–8 M forskolin) reduced the contractile response to AVP, while prostanoid synthesis (3 x 10–6 M indomethacin) inhibition and blockade of Ca2+-activated potassium channels (KCa) (10–3 M tetraethylammonium (TEA)) enhanced AVP contraction. Age, gender and smoking also modified the AVP response. Conclusion: Our findings suggest a role for AVP as a modulator of vascular tone in human IMA. The effect of AVP is dependent on prostanoids and Ca2+-activated K+ channels, so its dysfunction in pathophysiological cardiovascular processes could mean that AVP, among other factors, produces vasospasm in IMA grafts.

Key Words: Coronary artery bypass grafting • Vasopressin • Vasospasm • Internal mammary artery


   1. Introduction
Top
 Abstract
 1. Introduction
2. Patients and methods
 3. Results
 4. Discussion
 References
 
The established safety and long-term effectiveness of coronary artery bypass grafting (CABG) [1] make surgery the best treatment option for advanced coronary artery disease. The human internal mammary artery (IMA) has earned its recognition as the gold standard for use as the autogenous bypass graft in CABG on the basis of its excellent late patency and resistance to arteriosclerosis. However, in vivo and in vitro experiments have demonstrated its potent vasoconstriction response to a great variety of vasoactive agents and a 10% rate of IMA stenosis has been described in the absence of established predisposing factors. In the last 20 years, many research efforts have searched for ways to reduce the incidence of vasospasm in CABG. In effect, vasospasm may occur during harvesting, grafting or immediately following CABG, and could lead to early myocardial ischaemia and thus, increase perioperative morbidity and mortality in high-risk patients. Vasodilators such as calcium antagonists, long-acting nitrates and phosphodiesterase inhibitors are usually administered before grafting to increase IMA blood flow during surgery [2]. However, these agents seem to be more effective in reducing rather than preventing graft spasm [3]. One of the possible aetiologies of vasospasm is impaired endothelial-dependent blood flow caused by an inability of the endothelium to synthesize adequate amounts of nitric oxide (NO), evoking unopposed vasoconstriction [4]. Vasoconstriction (or spasm) may be produced by various, or a combination of, stimuli, such as surgical trauma, nerve stimulation and the release of several vasoconstrictor substances during CABG, which could include endothelin, tromboxane A2, prostaglandin F2{alpha}, extracellular nucleotides, serotonin, circulating sympathetic amines (norepinephrine and epinephrine), angiotensin II or vasopressin (AVP) [3]. In effect, it would be interesting to correlate the concentrations of these vasoconstrictors that could be released in the perioperative period, with the response shown by arterial grafts to these agents [5]. AVP, a hypothalamic hormone stored and released by the neurohypophysis, has marked vasoconstrictor effects and also has antidiuretic effects on the renal tubular system. The release of AVP is stimulated by conditions that are common in revascularization operations such as changes in systemic blood pressure, hypertonicity or hypovolaemia, amongst others factors. In cultured vascular smooth muscle cells, all the effects of AVP are mediated through the V1a receptor, which signals through G-proteins [6]. AVP increases intracellular Ca2+ via mobilization of intracellular stores and influx of extracellular Ca2+ via voltage-activated Ca2+channels. Increased intracellular Ca2+ also leads to increased arachidonic acid release and eicosanoid production through the action of phospholipase A2 [6].

To date, in vitro studies addressing the effects of AVP on the IMA have focused on AVP receptors [7] and the inhibitory effects of vasodilators on AVP-mediated contraction [8,9]. AVP has also recently gained attention as a possible tool against septic shock and vasodilator states associated with cardiac anaesthesia and surgery, although future prospective studies are necessary to define the role of AVP in the therapy of vasodilator shock [10].

In an effort to establish the real contribution of this neurohormone to vasospasm, the present study was designed to examine the in vitro vascular reactivity of IMA towards AVP after exposure to in vivo AVP release during CABG. The mechanisms involved in the AVP contraction were explored and we also tried to assess the effects of the main risk factors for cardiovascular disease on the AVP response.


   2. Patients and methods
Top
 Abstract
 1. Introduction
 2. Patients and methods
3. Results
 4. Discussion
 References
 
The study protocol was approved by the Ethics Committee of the Hospital General Universitario Gregorio Marañón, Madrid, Spain.

2.1 Anaesthesia and cardiopulmonary bypass management
Pharmacological therapy (platelet antiaggregants, ACE inhibitors and calcium channel blockers) was interrupted at least 1 week before CABG.

Anaesthesia was induced in all patients according to a standardized protocol including intravenous midazolam (0.03–0.05 mg/kg), remifentanil (1.0–3.0 µg/kg) and cifatracurium (0.5 mg/kg), and maintained with 0.2–0.5% isoflurane and continuous intravenous infusion of remifentanil at 0.1–0.5 µg/kg/min.

All patients underwent a routine single surgical technique. After gaining entry through a median sternotomy, the IMA was dissected simultaneously with the saphenous vein and/or the radial artery. In all patients, the IMA was harvested as a pedicle with the surrounding muscle and fascia, avoiding any external and/or internal pharmacological manipulation. Following systemic heparinization, the IMA was clipped distally, cut and occluded proximally with a bulldog clamp. IMA samples were harvested from the distal end, just before anastomosis, to the target coronary artery.

Cardiopulmonary bypass (CPB) was established by ascending aortic and two-stage right atrial cannulation (aortic cross-clamp time 50–60 min). Heparin was given at a dose of 300 IU/kg to achieve a target-activated coagulation time over 450 s. Moderate haemodilution (haematocrit 20–25%), mild hypothermia (30–32 °C) and a constant perfusion pressure (2.5 l/min/m2) were maintained during CPB (CPB time 90–100 min). The bypass circuit was primed with 750 ml of Ringer's lactate solution, 500 ml of hydroxy ethyl starch and 200 ml of 1/6 M sodium bicarbonate.

Myocardial arrest was induced through 1000 ml of antegrade and retrograde high-potassium (30 meq./l) cold (4 °C) blood cardioplegia (4:1 ratio). Thereafter, and for the remainder of the procedure, 500 ml low-potassium cardioplegia (10–15 meq./l) was administered through the coronary sinus at the end of each distal anastomosis. Reperfusion was controlled with warm low-potassium retrograde cardioplegia while performing proximal anastomoses. The mean intraoperative arterial pressure of the patients was 60–70 mmHg in all the stages of surgery.

2.2 Plasma AVP and serum osmolality, sodium and potassium
Blood samples were collected at the following stages into vacuum tubes from the arterial line of 16 patients subjected to routine CABG involving cardiopulmonary bypass:

1. Before induction of anaesthesia
2. After induction of anaesthesia and/or 10 min before CPB
3. Shortly after starting CPB
4. In the intensive care unit

Once collected, the blood samples were kept in ice slush, and plasma and serum separated by centrifugation (10 min, 4 °C) at 1620 x g. All samples were then frozen at –70 °C and stored until analysis.

Plasma AVP and serum osmolality, sodium and potassium were determined by the following methods:

AVP was determined using the radioimmunoassay kit Vasopressin Direct RIA (Buhlmann Laboratories AG, Switzerland), whose sensitivity is 1.2 pg/ml.

Blood osmolality was determined by the freezing-point osmometric method using an osmometer mod. 3D II (Advanced Instruments, USA). This procedure has a resolution of 50 mOsm/kg.

Serum sodium and potassium concentrations were measured by flame photometry using a photometer mod. 943 (Instrumentation Laboratory, USA). Sensitivities for sodium and potassium were 3 mmol/l and 0.1 mmol/l, respectively.

In the 16 patients in whom plasma AVP levels were determined, the postoperative course was uncomplicated, so there was no need for vasoactive drugs, with the exception of nitroglycerine, which was occasionally required to control arterial pressure.

2.3 Arterial rings
Artery segments were obtained from the portions of the IMA harvested for use as grafts during CABG surgery. The segments were placed in chilled, cold physiological saline solution (PSS) of composition (mM): NaCl, 119; KCl, 4.7; CaCl2, 1.5; MgSO4, 1.2; NaHCO3, 25; glucose, 10; KH2PO4, 1.2 and ethylene diaminetetraacetic acid (EDTA), 0.026, and transported to the laboratory. Once in the laboratory, adjacent tissue was carefully dissected away under a stereomicroscope (Nikon SMZ 2B, Japan) and each IMA segment was cut into rings, 2–3 mm in length, for isometric force recordings. The rings (external diameter = 1.35 ± 0.02 mm and internal diameter = 0.75 ± 0.01 mm; n = 201) were suspended on two parallel L-shaped stainless steel wires (diameter 150 µm) in 5 ml organ baths containing PSS at 37 °C and gassed with carbogen (95% O2 and 5% CO2) to maintain the pH at 7.4. One wire was fixed to a displacement unit allowing fine adjustment of tension while the other was attached to a force transducer (Grass FT03C, USA). The isometric tension of the artery wall was displayed and recorded using a PowerLab data acquisition system (PowerLab/8, model MLS013/W, AD Instruments Pty. Ltd., Australia). In some experiments, the endothelium was mechanically removed by careful rubbing of the inner surface of the segment with a stainless steel wire before mounting the segment.

2.4 Mechanical responses
After an equilibration period of 30 min in PSS, each ring was stretched in a stepwise fashion to the optimal point of its length–tension ratio ({approx}25 mN) determined in previous experiments. The arterial rings were allowed a 2-h normalization period before testing. In all the experiments, arterial specimens were exposed to a depolarizing potassium solution (124 mM, K-PSS) to evaluate the viability of the preparations. K-PSS was identical to PSS except that NaCl was replaced with KCl on an equimolar basis. The K-PSS response obtained was used as the reference contraction for each arterial ring. The integrity of the vascular endothelium was confirmed by relaxation induced by acetylcholine (ACh; 10–6 M) in rings precontracted with noradrenaline (NA; 10–6–3 x 10–6 M). This NA precontraction corresponded to 50–60% of the contraction induced by K-PSS. In experiments in which the endothelial layer was intentionally removed, ACh-induced relaxation was abolished while papaverine (10–4 M) relaxation was maintained. Each experimental procedure was performed on arterial specimens from at least six patients.

AVP was added to the organ baths in a cumulative fashion. Previous experiments showed that two consecutive cumulative concentration–response curves to AVP were not reproducible due to AVP-induced tachyphylaxis. Thus, it was necessary to use consecutive segments from the same patient in parallel experiments, with one ring acting as the control for the other, and only one AVP concentration–response curve per arterial ring was obtained.

To evaluate the effects on the AVP response, the different antagonists and/or blockers were incubated for 30 min before obtaining a concentration–response curve.

2.5 Drugs
The following drugs were used: acetylcholine hydrochloride, arginine vasopressin, catalase (radical scavenger), deferoxamine mesylate (inhibitor of hydroxyl radicals), forskolin (adenylate cyclase activator), indomethacin (prostanoid synthesis inhibitor), L-arginine hydrochloride (NO precursor), {omega}-nitro-L-arginine methyl ester hydrochloride (L-NAME; NO synthase inhibitor), noradrenaline hydrochloride, 1H-[1,2,4]oxadiazol [4,3,-{alpha}]quinoxalin-1-one (ODQ; soluble guanylyl cyclase blocker), papaverine hydrochloride (phosphodiesterase inhibitor), prazosin hydrochloride ({alpha}1-adrenergic receptor antagonist), superoxide dismutase (SOD; radical scavenger), tetraethylammonium (TEA; Ca2+-activated potassium channels (KCa) blockade) (all from Sigma Chemical Co., USA) and bosentan (non-specific endothelin receptor antagonist; a gift from Hoffmann-La Roche, Inc., USA).

All drugs were dissolved in distilled water with the following exceptions: indomethacin and papaverine were prepared in 96% ethanol, and ODQ and forskolin were dissolved in dimethyl sulphoxide and further diluted in water. Previous experiments showed that the solvents used had no effect on the preparations.

2.6 Data and statistical analysis
Plasma AVP and serum osmolality, sodium and potassium were compared with the Student's t-test for paired observations, using the values obtained in stage 1 (before anaesthesia) as controls. The level of significance was set at a P-value less than 5% (P < 0.05). Plasma AVP was compared to serum osmolality and serum sodium and potassium levels for each sampling period by the Pearson's correlation method. Maximal plasma AVP concentrations were also correlated with the maximal AVP contractions recorded in vitro for the same patient.

The tension of the vessel wall was measured in mN. Contractions obtained both on basal tone and in precontracted vessels were expressed as the percentage of contractions induced by K-PSS. Relaxations were expressed as the percentage of the vascular contraction induced by NA.

Sensitivity to AVP was expressed in terms of pEC50 values, where pEC50 = –log EC50, EC50 being the agonist concentration needed to produce a half-maximal response. pEC50 was estimated by computerized non-linear regression analysis (GraphPad Prism, USA).

Biochemical determinations and mechanical vascular responses were expressed as mean values ± standard error of the mean (SEM). Statistical analysis using the Student's t-test for paired and unpaired observations was performed when appropriate. For testing the effects of cardiovascular risk factors on the in vitro AVP response, the data were analysed by analysis of variance (ANOVA) and Duncan's post hoc comparison. In this analysis, a P-value less than 5% (P < 0.05) was also considered to denote significance.


   3. Results
Top
 Abstract
 1. Introduction
 2. Patients and methods
 3. Results
4. Discussion
 References
 
3.1 Patient characteristics
Segments of the IMA were obtained from 95 patients (mean age 64.2 ± 0.9 years): 80 men and 15 women. Their characteristics and risk factors for cardiovascular disease are shown in Fig. 1 .


Figure 1
View larger version (17K):
[in this window]
[in a new window]

  		Fig. 1. Characteristics and cardiovascular risk factors of the patients who underwent CABG in this study. BMI, body mass index; PMI, prior myocardial infarction.

 
3.2 Plasma AVP concentrations and variables measured
Circulating AVP levels recorded at stage 1 (before anaesthesia) were taken as baseline values for the statistical tests. These values were within physiological limits (≤7.6 pg/ml). At stage 2 (after anaesthesia induction but before CPB) AVP levels were 10-fold compared to baseline levels and continued to rise, peaking after the start of CPB (stage 3) and declining in the intensive-care unit after surgery (stage 4) (Table 1 ).


View this table:
[in this window]
[in a new window]

  		Table 1 Plasma AVP concentrations and variables measured
 
Among the remaining variables measured, only serum osmolality showed a significant increase, with similar sodium and potassium levels detected before, during and after CPB (Table 1). The elevated AVP plasma levels recorded in stage 3 correlated well with serum osmolality determined at the same sampling time for the same patient (Pearson's r = 0.9490; P < 0.0001; n = 16).

3.3 Vascular function
3.3.1 AVP response
Segments of human IMA contracted in a concentration-dependent manner in response to AVP (10–12 to 3 x 10–7 M). This neurohormone produced a long-lasting effect with a pEC50 value of 9.13 ± 0.09 (n = 24) and a maximal contraction (MC) of 42.6 ± 4.5% of K-PSS (2.52 ± 0.13 g; n = 24) (Fig. 2A).


Figure 2
View larger version (13K):
[in this window]
[in a new window]

  		Fig. 2. (A) Concentration–response curves for AVP in endothelium-intact and denuded preparations obtained on the basal tone of IMA. Each point represents the mean ± SEM of 24 experimental determinations. (B) Correlation between plasma AVP levels (pg/ml) recorded at stage 3 (after starting CPB) and the maximal contraction induced by AVP in vitro in IMA rings from the same patients (y = –15.468x + 740.55; Pearson's r = 0.6968; P < 0.01; n = 16).

 
The lack of a relaxant effect of AVP was demonstrated in vessel rings contracted with NA (10–6 to 3 x 10–6 M). In these conditions, AVP also exclusively evoked a contractile effect (pEC50 = 8.63 ± 0.16 and MC = 30.7 ± 5.4%; n = 13).

The possible role of contractile {alpha}1-adrenergic receptors was ruled out, since no effect on AVP action was shown by the specific {alpha}1-receptor antagonist (pEC50 = 9.30 ± 0.11 and MC = 41.9 ± 17.7% for controls; pEC50 = 8.92 ± 0.20 and MC = 36.3 ± 7.2% for 10–6 M prazosin; n = 7). Papaverine (10–4 M) caused full relaxation of the AVP-elicited contraction within approximately 25 min (n = 11).

An inverse correlation was observed between plasma AVP levels recorded at stage 3 (patient under anaesthesia shortly after starting CPB) and the maximal contraction induced by AVP in vitro in vascular rings from the same patients (Pearson's r = –0.6968; P < 0.01; n = 16) (Fig. 2B).

3.3.2 Effects of endothelial denudation, NO synthase and guanylate cyclase inhibition, prostanoid synthesis and KCa blockade on basal tone and AVP-induced contraction
Mechanical removal of the endothelium failed to modify both basal tension and the contraction response to AVP, which was similar in arterial rings with an intact endothelium (pEC50 = 9.33 ± 0.32 and MC = 57.1 ± 12.9%) and denuded rings (pEC50 = 9.28 ± 0.24 and MC = 54.1 ± 12.2%; n = 11) (Fig. 2A).

The following experiments were, nevertheless, performed to determine whether the AVP response could be modulated by the release of endothelium-derived relaxant and contractile factors in a delicate balance. Inhibition of NO synthase with L-NAME (3 x 10–5 M) produced a discrete 4.7 ± 0.9% rise in basal tension with no significant displacement of the AVP concentration–response curve (pEC50 = 8.98 ± 0.14 and MC = 51.9 ± 6.7% for controls; pEC50 = 9.18 ± 0.18 and MC = 63.6 ± 7.8% for L-NAME; n = 13). The NO precursor, L-arginine (3 x 10–4 M), failed to decrease the AVP contraction response after incubation periods of both 30 min (pEC50 = 8.94 ± 0.13 and MC = 63.7 ± 11.8% for controls; pEC50 = 9.26 ± 0.16 and MC = 57.4 ± 7.9% for 3 x 10–4 M L-arginine; n = 11) and 8 h (pEC50 = 8.92 ± 0.10 and MC = 42.0 ± 1.7% for controls; pEC50 = 9.00 ± 0.08 and MC = 39.2 ± 4.5% for L-arginine; n = 8).

In intact and denuded arterial rings, blockade of cyclooxygenase with indomethacin (3 x 10–6 M) evoked a moderate increase in basal tone (17.0 ± 2.2%, n = 7 and 19.2 ± 2.8%; n = 6, respectively). However, the simultaneous addition of L-NAME (3 x 10–5 M) and indomethacin (3 x 10–6 M) led to a markedly enhanced basal tone both in intact (70.8 ± 14.3%; n = 9; P < 0.01) and denuded arterial rings (60.0 ± 19.5%; n = 8; P < 0.05).

Indomethacin (3 x 10–6 M) produced a moderate yet significant increase in the sensitivity of the AVP concentration–response curve (Fig. 3A). The adenylate cyclase activator, forskolin (3 x 10–8 M), inhibited the contractile response to AVP (Fig. 3A). Simultaneous pre-treatment with L-NAME (3 x 10–5 M) and indomethacin (3 x 10–6 M) significantly enhanced the sensitivity and maximal contractile response to AVP (Fig. 3B). Indomethacin (3 x 10–6 M) plus ODQ (3 x 10–6 M), however, did not modify the contractions to AVP (Fig. 3B).


Figure 3
View larger version (18K):
[in this window]
[in a new window]

  		Fig. 3. (A) Concentration–response curves for AVP in the absence/presence of 3 x 10–6 M indomethacin and the absence/presence of 3 x 10–8 M forskolin recorded in rings of IMA. Each point represents the mean ± SEM of 6–8 experimental determinations, respectively. (B) Concentration–response curves for AVP in the absence/presence of 3 x 10–6 M indomethacin plus 3 x 10–5 M L-NAME and in the presence of 3 x 10–6 M indomethacin plus 3 x 10–6 M ODQ recorded in rings of IMA. Each point represents the mean ± SEM of 7–11 experimental determinations, respectively.

 
When KCa channels were blocked with TEA (10–3 M) a significant increase in the maximal response to AVP was produced with no effects on resting tension (Fig. 4 ).


Figure 4
View larger version (10K):
[in this window]
[in a new window]

  		Fig. 4. Concentration–response curves for AVP in the absence/presence of 10–3 M TEA recorded in rings of IMA. Each point represents the mean ± SEM of 6 experimental determinations.

 
3.3.3 Effects of the removal of superoxide anions and hydroxyl radicals and endothelin blockade on basal tone and AVP-induced contraction
To explore the role of endothelium-derived contractile factors, we added SOD (100 U/ml) plus catalase (1200 U/ml), deferoxamine (10–4 M) and bosentan (10–5 M) to the bath. SOD plus catalase mop up superoxide anions producing hydrogen peroxide. Deferoxamine inhibits hydroxyl radical synthesis. Bosentan is a non-specific endothelin receptor antagonist. However, none of these substances had any effects on basal tone or AVP-induced contraction (Table 2 ).


View this table:
[in this window]
[in a new window]

  		Table 2 Effects of SOD + catalase, deferoxamine or bosentan on AVP-induced contraction
 
3.4 Effects of cardiovascular risk factors on AVP-induced contraction
Among all the risk factors analysed, the maximal contraction induced by AVP in the IMA rings varied significantly according to age, gender and cigarette smoking (Table 3 ). AVP-induced contraction was higher in patients older than 65 years compared to the younger patients, while a significantly lower response to AVP was observed in men and in smokers (Table 3).


View this table:
[in this window]
[in a new window]

  		Table 3 Maximal contraction produced in response to AVP recorded in IMA specimens from patients with or without risk factors for cardiovascular disease
 

   4. Discussion
Top
 Abstract
 1. Introduction
 2. Patients and methods
 3. Results
 4. Discussion
References
 
The findings of our study indicate that: (a) the contractile response of the human IMA to AVP is endothelium-independent and involves smooth muscle prostanoids and KCa channels; (b) maximal AVP-induced vasoconstriction of IMA in vitro is inversely related to the plasma AVP concentration and (c) age, gender and smoking seem to affect the IMA response to AVP.

The AVP hormone is essential for osmotic and cardiovascular homeostasis. Given that AVP could provoke graft spasm [8], we examined the release of AVP before, during and after CABG surgery, and assessed ways to prevent its contractile effect.

An increase in plasma levels of AVP was observed during CABG that was most pronounced at the onset of CPB, in agreement with the previous findings [11]. AVP release is thought to be mainly influenced by the activity of osmoreceptors, volume receptors, baroreceptors, and pain and visceral sensory input [11]. The high level of osmolality observed here just after the start of CPB and after surgery and its correlation with AVP plasma concentrations suggest that osmoreceptor activation could be one of the key triggers for elevating plasma AVP levels. A mechanism involving volume receptors would also likely be induced by the lowered left atrial pressure during extracorporeal circulation, although AVP is more sensitive to small osmolality changes than equivalent variations in plasma volume [11]. Given that the mean perfusion pressure did not change during CPB, the activation of baroreceptors by hypotension is unlikely. Neither does the involvement of a mechanism related to pain and visceral sensory input, which is known to release AVP, seems plausible, since the AVP increase started after inducing anaesthesia, when there would be no sensory input. The surge of AVP detected during and after CPB could therefore promote the undesirable effect of reducing organ perfusion [11]. Our findings indicate that, through the mechanisms cited, CPB seems to be associated with the AVP surge recorded here in patients undergoing CABG. Comparable levels of plasma AVP at the end of surgery have been reported by Velissaris et al. [12]. These authors, however, examined hormone release related to perioperative stress in patients undergoing CABG with and without CPB, and did not assess AVP changes during the CPB process.

AVP is regarded as being among the most potent vasoconstrictors in the organism in terms of both the maximal contractile effect induced and the duration of this effect. Our results reveal that AVP provokes a slow-onset, long-lasting in vitro contraction persisting for up to 2 h. Today, V1a is established as the subtype of AVP receptor involved in vascular contraction [10] and it is this receptor that displays rapid tachyphylaxis [13]. Hence, the desensitization of IMA towards AVP noted in the present study indirectly indicates that V1a is the receptor involved in the AVP response.

Moreover, the rapid desensitization of IMA to AVP could explain the inverse correlation we observed between plasma AVP and the maximal contraction induced by AVP in vitro. Desensitization is a complex process that plays an important role in turning off receptor-mediated signal transduction pathways. Signal transduction pathways that turn on also need to be turned off. This ensures that signalling occurs in a spatiotemporal manner so that cell function can be finely regulated [14]. Some authors suggest that V1a receptors are downregulated when blood concentrations of AVP are high, which would explain the decreased sensitivity to the vascular effects of AVP observed in induced sepsis [15] and in transgenic rats overexpressing AVP [16]. These findings suggest there may be a protective mechanism against AVP elevation to maintain normal cardiovascular regulation. However, the precise mechanism responsible for this remains unclear.

Vascular smooth muscle relaxation is related to the biological activity of second messengers such as cyclic adenosine monophosphate and cyclic guanosine monophosphate, which are hydrolysed by phosphodiesterases. Papaverine is a phosphodiesterase inhibitor widely used in patients undergoing CABG since it seems to be effective in preventing graft spasm. In our experiments, papaverine reversed the AVP-induced contraction in every vessel ring tested. This agent could thus serve to suppress any vasospasm induced by AVP, although its detrimental effects on vascular endothelial function would need to be considered [17].

The contraction response of the IMA to AVP is mediated by its direct action on vascular smooth muscle. The role of the endothelium in regulating the AVP contractile effect in the human IMA was ruled out here since removal of the endothelium had no effect. Our detailed analysis of endothelial relaxant and contractile factors confirmed this result. Thus, the endothelium-derived relaxant, NO, did not seem to be involved in the AVP response since L-NAME failed to modify the AVP concentration–response curve. However, the non-selective COX inhibitor, indomethacin, was found to significantly enhance the AVP contractile effect both in intact and denuded arterial specimens, indicating the release of relaxant prostanoids from the smooth muscle layer after AVP binds to its specific receptor. Indomethacin plus L-NAME dramatically increased their response to AVP in endothelium-denuded segments. These results suggest cross-talk regulation between prostanoids and nitric oxide in the smooth muscle cells of the arterial wall, and could explain the NO-synthase activity observed when the cyclooxygenase pathway is inhibited. Illiano et al. [18] described this mechanism in the regulation of basal tone in canine saphenous vein segments devoid of endothelium. These authors suggest that the basal production of prostanoids by smooth muscle cells could be sufficient to trigger the activation of cyclic AMP-dependent protein kinase, which in turn could catalyse the downregulation of nitric oxide synthase. In the present study, we also noted that indomethacin plus L-NAME increased basal tone such that this could also be the mechanism underlying the AVP-induced supra-maximal contraction in the IMA. Moreover, interaction between the two signal transduction pathways could occur at the NO synthesis enzyme level because treatment with indomethacin plus ODQ caused no further enhancement of the AVP response beyond that observed when indomethacin was added separately.

In addition to these mechanisms, KCa channels seem to attenuate AVP contraction since blockade with TEA increased the contractions to AVP. K+ channels induce the relaxation or reduce the contraction of blood vessels through hyperpolarization of vascular smooth muscle cells and reducing the concentration of cytoplasmic Ca2+. AVP increases intracellular Ca2+ by mobilizing intracellular stores and allowing the influx of extracellular Ca2+. Hence, KCa channels may, through negative feedback, limit active vasoconstriction induced by AVP. This potentiation effect of KCa channel blockade on AVP contraction has also been observed in the renal arteries of male rats [19].

KCa channels may compensate for the loss of other vasodilatory mechanisms in disease states in which contractile factor levels are elevated. In effect, increased levels of superoxide radicals have been observed in cardiovascular disorders. Although these radicals may activate Ca2+-activated K+ channels [20], this is probably not the activation mechanism of these channels in the IMA, since the superoxide scavengers, SOD plus catalase, did not modify the AVP contraction. Further, we were able to rule out the participation of another endothelium-derived contractile factor since deferoxamine and bosentan did neither reduce the basal tone nor compromise the AVP contractile response.

Among all the risk factors examined, the maximal contraction induced by AVP in the rings of IMA was significantly modified by age, gender and cigarette smoking. Several studies have revealed an increased effect of vascular constrictors with age in agreement with the greater response recorded in arterial specimens corresponding to the eldest patients included in our study. This finding could be attributable to the reduced release of NO or EDHF as vasodilators, or the increased release of vasoconstrictor prostaglandins. Although the mechanism of AVP-induced vascular hyperreactivity remains to be elucidated, due to its endothelial-independent nature, Ca2+ and/or K+ channels located in smooth muscle could be involved.

Nicotine stimulation induced by cigarette smoking has previously been identified as a potent stimulus for AVP release in humans [21]. Our results indicate a reduced contractile response to AVP in smokers. The higher circulating plasma AVP levels in this patient group may contribute to the in vivo AVP receptor desensitization mechanisms mentioned above, possibly translating to a reduced vascular response to AVP in vitro. Another possible explanation could be that since many people seem to give up smoking as they become older, the group of smokers was of a younger mean age.

It is well established that the risk of developing coronary artery disease and hypertension is much higher (3–4 times) in men than in premenopausal women, whereas in women, after the menopause, this gender difference is reversed. The women in our study were postmenopausal and AVP showed a more pronounced response in women than in men. An inhibitory role of oestrogens in AVP's actions has been suggested [22], which would be consistent with the present results. Notwithstanding, numerous clinical and epidemiologic studies describe a controversial relationship between gender and cardiovascular disease, as well as between gender and vascular reactivity [23]. Li and Stallone [24] recently established that oestrogen potentiates AVP-induced contraction in the aorta of the female rat by enhancing cyclooxygenase-2 and tromboxane function. Sex hormones may be involved in this gender difference, although we cannot rule out the possible effects of age on women vasopressinergic vasoconstriction [25].

In conclusion, our findings suggest a role for AVP as a modulator of vascular tone in human IMA. The effect of AVP is dependent on prostanoids and Ca2+-activated K+ channels, so its dysfunction in pathophysiological cardiovascular processes could mean that AVP is responsible, among other factors, for the vasospasm observed in IMA grafts.


   Acknowledgments
 
The authors would like to thank the surgeons and nurses of the heart surgery unit of the Hospital General Universitario Gregorio Marañón (Madrid, Spain) for their help in the operating room.


   Footnotes
 
\#9734; This study was supported by grant PI 031257 from the FIS, Spanish Ministry of Health.


   References
Top
 Abstract
 1. Introduction
 2. Patients and methods
 3. Results
 4. Discussion
 References
 

  1. Taggart DP. Surgery is the best intervention for severe coronary artery disease. BMJ 2005;330:785-786.[Free Full Text]
  2. Takeuchi K, Sakamoto S, Nagayoshi Y, Nishizawa H, Matsubara J. Reactivity of the human internal thoracic artery to vasodilators in coronary artery bypass grafting. Eur J Cardiothorac Surg 2004;26(5):956-959.[Abstract/Free Full Text]
  3. Rosenfeldt FL, He GW, Buxton BF, Angus JA. Pharmacology of coronary artery bypass grafts. Ann Thorac Surg 1999;67(3):878-888.[Abstract/Free Full Text]
  4. He GW. Nitric oxide and endothelium-derived hyperpolarizing factor in human arteries and veins. J Card Surg 2002;17(4):317-323.[Medline]
  5. Lockowandt U, Bjessmo S, Ivert T, Franco-Cereceda A. Plasma levels and vascular effects of endothelin and big endothelin in patients with stable and unstable angina pectoris undergoing coronary bypass grafting. Eur J Cardiothorac Surg. 2002;21(2):218-223.[Abstract/Free Full Text]
  6. Nemenoff RA. Vasopressin signaling pathways in vascular smooth muscle. Front Biosci 1998;15(3):194-197.
  7. Burrell LM, Phillips PA, Rolls KA, Buxton BF, Johnston CI, Liu JJ. Vascular responses to AVP antagonists in man and rat. Clin Sci (Lond) 1994;87(4):389-395.[Medline]
  8. Wei W, Floten HS, He GW. Interaction between vasodilators and vasopressin in internal mammary artery and clinical significance. Ann Thorac Surg 2002;73(2):516-522.[Abstract/Free Full Text]
  9. Wei W, Yang CQ, Furnary A, He GW. Greater vasopressin-induced vasoconstriction and inferior effects of nitrovasodilators and milrinone in the radial artery than in the internal thoracic artery. J Thorac Cardiovasc Surg 2005;129(1):33-40.[Abstract/Free Full Text]
  10. den Ouden DT, Meinders AE. Vasopressin: physiology and clinical use in patients with vasodilatory shock: a review. Neth J Med 2005;63(1):4-13.[Medline]
  11. Wu W, Zbuzek VK, Bellevue C. Vasopressin release during cardiac operation. J Thorac Cardiovasc Surg 1980;79(1):83-90.[Medline]
  12. Velissaris T, Tang AT, Murray M, Mehta RL, Wood PJ, Hett DA, Ohri SK. A prospective randomized study to evaluate stress response during beating-heart and conventional coronary revascularization. Ann Thorac Surg 2004;78(2):506-512.[Abstract/Free Full Text]
  13. Hamel C, Millette E, Lamontagne D. Role of nitric oxide and protein kinase C in the tachyphylaxis to vasopressin in rat aortic rings. Life Sci 2005;77(10):1069-1081.[CrossRef][Medline]
  14. Bünemann M, Lee KB, Pals-Rylaarsdam R, Roseberry AG, Hosey MM. Desensitization of G-protein-coupled receptors in the cardiovascular system. Annu Rev Physiol 1999;61:169-192.[CrossRef][Medline]
  15. Bucher M, Hobbhahn J, Taeger K, Kurtz A. Cytokine-mediated downregulation of vasopressin V(1A) receptors during acute endotoxemia in rats. Am J Physiol Regul Integr Comp Physiol 2002;282(4):R979-R984.[Abstract/Free Full Text]
  16. Tachikawa K, Yokoi H, Nagasaki H, Arima H, Murase T, Sugimura Y, Miura Y, Hirabayashi M, Oiso Y. Altered cardiovascular regulation in arginine vasopressin-overexpressing transgenic rats. Am J Physiol Endocrinol Metab 2003;285(6):E1161-E1166.[Abstract/Free Full Text]
  17. Gao YJ, Yang H, Teoh K, Lee RM. Detrimental effects of papaverine on the human internal thoracic artery. J Thorac Cardiovasc Surg 2003;126(1):179-185.[Abstract/Free Full Text]
  18. Illiano S, Marsault R, Descombes JJ, Verbeuren T, Vanhoutte PM. Regulation of nitric oxide-like activity by prostanoids in smooth muscle of the canine saphenous vein. Br J Pharmacol 1996;117(2):360-364.
  19. Sanz E, Fernandez N, Monge L, Climent B, Dieguez G, Garcia-Villalon AL. Role of K(+) channels in the coronary and renal vascular reactivity to vasopressin in diabetic rats. Eur J Pharmacol 2003;471(1):35-40.[CrossRef][Medline]
  20. Liu Y, Gutterman DD. The coronary circulation in diabetes: influence of reactive oxygen species on K(+) channel-mediated vasodilation. Vascul Pharmacol 2002;38(1):43-49.[CrossRef][Medline]
  21. Pomerleau OF. Nicotine and the central nervous system: biobehavioral effects of cigarette smoking. Am J Med 1992;93(1A):2S-7S.[CrossRef][Medline]
  22. Ishunina TA, Kruijver FP, Balesar R, Swaab DF. Differential expression of estrogen receptor alpha and beta immunoreactivity in the human supraoptic nucleus in relation to sex and aging. J Clin Endocrinol Metab 2000;85(9):3283-3291.[Abstract/Free Full Text]
  23. Sader MA, Celermajer DS. Endothelial function, vascular reactivity and gender differences in the cardiovascular system. Cardiovasc Res 2002;53:597-604.[Free Full Text]
  24. Li M, Stallone JN. Estrogen potentiates vasopressin-induced contraction of female rat aorta by enhancing cyclooxygenase-2 and thromboxane function. Am J Physiol Heart Circ Physiol 2005;289(4):H1542-H1550.[Abstract/Free Full Text]
  25. Loichot C, Anjuere J, Nisato D, De Jong W, Imbs JL, Barthelmebs M. Renal vascular reactivity to vasopressin in rats with diabetes mellitus. Eur J Pharmacol 2001;431(3):321-329.[CrossRef][Medline]




This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to Personal Folders
Right arrow Download to citation manager
Right arrow Permission Requests
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Novella, S.
Right arrow Articles by Benedito, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Novella, S.
Right arrow Articles by Benedito, S.
Related Collections
Right arrow Cardiac - pharmacology
Right arrow Coronary disease
Right arrow Extracorporeal circulation
Right arrow Great vessels

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
ANN THORAC SURG ASIAN CARDIOVASC THORAC ANN EUR J CARDIOTHORAC SURG
J THORAC CARDIOVASC SURG ICVTS ALL CTSNet JOURNALS