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Am J Physiol Heart Circ Physiol 284: H385-H392, 2003. First published September 19, 2002; doi:10.1152/ajpheart.00658.2002
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Vol. 284, Issue 1, H385-H392, January 2003

Systemic alpha 1A-adrenoceptor antagonist inhibits neointimal growth after balloon injury of rat carotid artery

John C. Teeters*, Cauveh Erami*, Hua Zhang, and James E. Faber

Department of Cell and Molecular Physiology, University of North Carolina, Chapel Hill, North Carolina, 27599-7545


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Previous in vitro and in vivo studies have shown that norepinephrine, acting through alpha 1A-adrenoceptors, stimulates hypertrophy, proliferation, and migration of vascular smooth muscle cells and adventitial fibroblasts and may contribute to neointimal growth, lumen loss, and inward remodeling caused by iatrogenic wall injury and vascular disease. Our present aim was to determine whether intravenous administration of the alpha 1A-adrenoceptor antagonist KMD-3213, at dosages without systemic hemodynamic effects, inhibits wall growth after injury. Inhibition of alpha 1A-adrenoceptors with 12.8 and 32 µg/kg KMD-3213 had no effect on arterial pressure or renal and hindquarter resistances in anesthetized rats. A second group then received carotid balloon injury and continuous intravenous KMD-3213 at 4 and 10 µg · kg-1 · h-1 for 2 wk. Mean, systolic, and diastolic arterial pressures and heart rate of conscious unrestrained rats were unaffected. KMD-3213 reduced neointima growth by ~30 and 46% at the two doses (P < 0.01). These data support the novel hypothesis that a direct alpha 1A-adrenoceptor-dependent trophic action of catecholamines is augmented by injury and may contribute significantly to hypertrophic vascular disease.

adrenergic receptor; smooth muscle cell


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

CERTAIN VASOACTIVE MEDIATORS are now recognized to contribute to the progression of hypertrophic vascular disease, in part through stimulatory (e.g., angiotensin) or loss of inhibitory (e.g., nitric oxide) effects on proliferation and hypertrophy of vascular smooth muscle cells (SMCs) and adventitial fibroblasts (AFBs) and the extracellular matrix they elaborate (4, 28). These effects derive from direct as well as indirect effects secondary to changes in pressure-dependent wall tension. There is evidence that the sympathetic neurotransmitter and hormone norepinephrine (NE) may also have direct trophic effects, although much less is known about this possibility. Most large arteries have an adrenergic plexus (2) commonly assumed to subserve reflex decreases in wall compliance during sympathoexcitation. However, studies using systemic chemical and immunological sympathetic denervation (14), chronic infusion of catecholamines (5, 18), or alpha -adrenoceptor (alpha -AR) antagonists (17), as well as correlation of plasma catecholamine levels in humans with wall hypertrophy and stiffness (6) and severity of atherosclerosis (19), suggest that NE released from these nerves may exert direct trophic effects. In animal models employing balloon injury of the carotid or aorta, chronic administration of nonsubtype-selective alpha 1-AR antagonists reduces neointimal growth by at least 50% (11, 16, 24, 34). However, potential effects of the concomitant hemodynamic disturbances on wall growth complicate interpretation of past in vivo studies regarding a direct trophic effect of NE. For example, systemic sympathetic denervation and chronic infusion of nonspecific alpha 1-AR antagonists cause hypotension that reduces wall tension, inhibits growth of SMCs (and possibly AFBs), and reduces extracellular matrix.

Recent studies (7, 10, 13, 25), however, do support a direct trophic action of catecholamines. Most arteries express multiple alpha -AR subtypes, some of which do not mediate vasoregulation and whose function has thus been unknown. For example, whereas contraction of the rat carotid and aorta is mediated by the alpha 1D-AR (7, 10, 13, 25), SMCs in the media of these vessels also express alpha 1A-, alpha 1B-, and alpha 2D-ARs (10). Moreover, AFBs express these same four alpha -ARs, and total alpha 1-AR density in adventitia is the same as that in the medial layer, although AFBs do not contract (10). beta -Adrenoceptors are also present on both cell types of these arteries (see Ref. 10 for references). It is well known that NE induces proliferation (14, 33, 36) and hypertrophy (26, 36) of cultured SMCs. We have recently found that NE also induces proliferation of cultured AFBs (10) and migration of both SMCs and AFBs that are mediated by different alpha -ARs (38). In rat aortas maintained in organ culture under wall tension, chronic NE exposure caused SMC hypertrophy and AFB proliferation and reduced SMC expression of proteins that characterize the contractile phenotype (39). These effects of NE were strongly augmented in aortas that had received balloon injury either 4 or 12 days earlier in vivo. Moreover, the effects of NE were blocked in intima-media cells by the alpha 1A-AR antagonist KMD-3213 and in adventitial cells by an alpha 1B-AR antagonist, whereas alpha 1D-, alpha 2-, and beta -AR antagonists had little or no effect (39).

Chronic local perivascular suffusion of NE and AR antagonists around normal and balloon-injured rat carotids (to avoid systemic hemodynamic effects) also supports the concept that wall NE exerts a direct trophic effect (8). In the uninjured artery, blockade of basal NE activity with alpha 1-AR antagonists had no effect on wall dimensions. However, elevating wall NE reduced circumference and lumen area (i.e., promoted eutrophic inward remodeling). In the injured artery, elevated wall NE had similar effects and, moreover, augmented neointimal area and collagen content, resulting in ~50% greater lumen loss. Furthermore, nonsubtype-specific alpha 1-AR antagonists and the alpha 1A-AR antagonist KMD-3213 reduced neointimal area by 33-54% and lumen loss by 50-70%, whereas alpha 1B- and alpha 1D-AR, alpha 2-, and beta -AR antagonists were without effect (8). These in vivo results suggest that elevated NE can act directly to cause adaptive wall hypertrophy in the uninjured artery and, moreover, that these trophic effects are augmented by injury such that even basal NE levels may contribute significantly to intimal expansion and lumen loss in injury-associated vascular disease. However, these results are limited by the potential effect of the perivascular infusion method itself, i.e., foreign body reaction (which, however, had no effect on the injury response) and inability to know tissue drug concentration. Therefore, it is important to test this novel hypothesis in vivo in the injured carotid artery with intravenous administration of an alpha 1A-AR antagonist at levels free of effects on arterial pressures and thus loading conditions on the vascular wall. The strategy is made feasible by the observation that the alpha 1D-AR mediates rat carotid contraction and is the major subtype mediating the sympathetic component of basal systemic vascular resistance in the rat (3, 7, 32).


    MATERIALS AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

KMD-3213 dose formulation. The Ki for KMD-3213 (alpha 1A-AR antagonist; kindly provided by Dr. Y. Kurashina, Kissei Pharmaceutical, Matsumoto City, Japan), determined for native (submandibular gland) and cloned rat alpha 1A-AR, averaged 0.28, and KMD-3213 showed 56- and 583-fold selectivity against alpha 1D- and alpha 1B-ARs, respectively (1, 37). This is the highest selectivity of alpha 1A-AR antagonist available. Because the pharmacokinetics of KMD-3213 are proprietary, several assumptions were required to determine dosage using standard pharmacokinetic equations (20). The IC50 for KMD-3213 against increases in rat prostatic intraurethral pressure (alpha 1A-AR dependent) evoked by intravenous phenylephrine (PE) is 0.12 mg/kg (1). Ten minutes after intravenous administration of the same doses, prazosin and KMD-3213 achieve near-equal plasma concentrations. This suggests that KMD-3213 and prazosin, which have similar solubilities and thus assumed bioavailabilities by an intravenous route, have similar volumes of distribution (0.6 l/kg) and plasma clearance rates and half-lives (0.18 l · kg-1 · h-1, 12 h) (20, 37). For the acute experiment (below), 12.8 µg/kg (low dose) was thus calculated to achieve the plasma concentration obtained by Akiyama et al. (1) (estimated to be 4 × 10-8 M); a 2.5-fold high dose (32 µg/kg) was also tested. These concentrations are predicted to selectively block 50-70% of the in vivo functional response of tissue (e.g., prostate) alpha 1A-ARs.

Acute experiment: effect of KMD-3213 on renal and hindquarter resistances, blood pressure, and PE sensitivity. To confirm the selectivity of KMD-3213, male Sprague-Dawley rats (n = 7, 300-400 g) were anesthetized with phenobarbital (60 mg/kg ip, with 10 mg/kg iv supplements given as needed) and received atropine (54 µg/kg sc). Rectal temperature was maintained with a heating pad. The abdominal aorta and inferior vena cava were catheterized via a femoral triangular incision. After laparotomy, a Doppler flow probe was placed around the left renal artery and suprailiac abdominal aorta. Wounds were closed, and skin edges were treated with lidocaine gel. A PE dose-response curve (1.0, 3.0, 5.0, and 10.0 µg/kg iv) was obtained, with doses separated by at least 3 min for recovery of baseline. Doses of PE were repeated 30 min after the administration of the low and then high doses of KMD-3213. Effects of KMD-3213 on baselines were measured during the last 5 min of both 30-min intervals. Peak responses to PE were compared with the control value for each parameter obtained during the 1-min interval immediately preceding drug administration.

Chronic experiment: systemic KMD-3213 infusion and balloon injury. Male 500-g Sprague-Dawley rats (n = 27) were anesthetized with ketamine (125 mg/kg im) plus acepromazine (1.25 mg/kg im) and received atropine (54 µg/kg sc) and cephazolin (160 mg/kg im). With the use of sterile surgery, the abdominal aorta was catheterized via the femoral artery, and the catheter (filled with 10 U/ml heparinized saline) was exposed at the back of the neck after subcutaneous anchoring with a Velcro "washer." The abdominal vena cava was catheterized via the femoral vein and connected to a primed osmotic minipump (5 µl/h; model 2ML2; Alza Durect, Cupertino, CA), which was placed into the abdomen via an inguinal incision. The venous catheter had four 26-gauge needle holes placed in the walls just above the end, which was sealed to prevent occlusion of the tip with refluxed blood. Pumps were filled with one of the following sterile, freshly prepared drugs: vehicle (50% ethanol in water), low-dose KMD-3213 (to give 4 µg · kg-1 · h-1), or high-dose KMD-3213 (to give 10 µg · kg-1 · h-1). KMD-3213 concentrations were determined with constant infusion equations (20) to achieve the plasma concentrations targeted in the acute experiment of ~4 × 10-8 M by the low dose and 1 × 10-7 M by the high dose.

The left thyroid, occipital, and external carotid arteries proximal to the lingual artery were ligated. The common carotid artery and overlying tissues were not disturbed below the carotid bifurcation. After heparin administration (63 U/kg im and 113 U/kg sc), a 2-Fr embolectomy catheter (Baxter Healthcare; Irvine, CA) was advanced via an external carotid arteriotomy to the origin of the left common carotid artery. The balloon was inflated with 15 µl of saline and rotated while it was withdrawn the length of the common carotid artery; the procedure was repeated twice, and the external carotid artery was ligated. The sham procedure consisted of exposure of the vessels above the right carotid bifurcation. All wounds were treated with nitrofurazone. Pentazocine (10 mg/kg im) was given for analgesia. Animals receiving KMD-3213 by osmotic pump were given a loading dose of 30 µg/kg to facilitate rapid achievement of the above steady-state concentrations. Procedures followed institutional guidelines. The effects of systemic KMD-3213 on the uninjured carotid artery were not examined, because, in previous studies, local perivascular administration of nonsubtype-specific alpha 1-AR antagonists, although reducing experimental restenosis, had no effect on dimensions of the uninjured carotid artery (8).

Blood pressure measurement. Beginning on the second postoperative day, hemodynamic data were collected on alternate days in conscious unrestrained rats in their home cage in a quiet, dimly lit room. The arterial catheter was flushed with 0.2 ml of heparinized saline (10 U/ml) and connected to a pressure transducer and chart recorder. Systolic, diastolic, and mean arterial pressures and heart rate were averaged over a 5-min interval after 30 min had passed, during which the animals rested quietly. Because patency of catheters declined during the second week, on postoperative day 14 animals were anesthetized with phenobarbital (60 mg/kg ip) and the femoral artery was catheterized to obtain final pressure determinations.

Morphometry. Pumps were verified as empty and their catheters as intact, and the vasculature was perfusion fixed at 100 mmHg with transcardial 4% paraformaldehyde in PBS (PFA). Left and right carotids were removed en bloc. Vessels were postfixed for 24 h in 4% PFA at 4°C, and the central 5-mm section of the common carotid artery was blocked and embedded in paraffin. Eight micron sections were cut every 200 µm and stained with Masson's trichrome (10 sections per vessel). Three trichrome-stained sections approximately equally separated over the 2-mm central section of the carotid were selected for digital planimetry (Scion Image, National Institutes of Health) by three observers blinded for the treatment group. Areas were determined as follows: lumen area = (lumen circumference)2/4pi , neointimal area = area between the lumen and internal elastic lamina (IEL), medial area = area between the IEL and external elastic lamina (EEL), circumference = length of EEL, and adventitial area = area of the dense collagen-containing layer between the EEL and loose perivascular connective tissue. Thickness of neointima was calculated as [(circumference of IEL divide  2pi ) - (circumference of lumen divide  2pi )]; thickness of media and adventitia was similarly calculated using circumferences of the IEL and EEL, and the outer edge of the dense adventitia and EEL, respectively.

Data are given as means ± SE for the number of vessels (1 per animal). Differences were subjected to unpaired t-tests or ANOVA followed by Bonferroni corrected t-tests for multiple comparisons (two-tailed unless indicated differently). A value of P < 0.05 was considered significant.


    RESULTS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Selectivity of KMD-3213, at the low and high doses, was determined for use in the subsequent chronic experiment (see Chronic experiment: systemic KMD-3213 infusion and balloon injury) during anesthesia with phenobarbital, which favors no change or a modest increase in sympathetic activity. Peripheral resistance under resting conditions is known to rely predominantly on alpha 1D-ARs in the rat (3, 32). Sympathetic constriction in the rat kidney, where alpha 1D-AR mRNA and binding sites are essentially undetectable, appears to be mediated by alpha 1B- and alpha 1A-ARs (29, 30), whereas data suggest that hindquarter (predominantly skeletal muscle) constriction may be mediated by alpha 1D- and alpha 1A-ARs (21, 26, 33). It is also known in the conscious as well as barbiturate-anesthetized rat that adrenergic contribution to basal resistance is relatively low in the kidney and high in the hindquarters (39). Neither low nor high doses of KMD-3213 altered baseline mean arterial pressure, renal resistance, or hindquarter resistance (Fig. 1). To examine the selectivity of KMD-3213 during increased catecholamine activity, we also examined the effect of KMD-3213 on pressor and constrictor responsiveness to PE. KMD-3213 had no significant effect on increases in arterial pressure (~30-40 mmHg) and renal and hindquarter resistances produced by low PE doses (0.5 and 1 µg/kg; Fig. 1). At high PE doses that were, in the absence of KMD-3213, supramaximal for increases in arterial pressure, KMD-3213 inhibited the pressor and resistance increases in the kidney and hindquarters (Fig. 1). This suggests that alpha 1A-ARs contribute to increased vascular resistance in these beds during sympathoexcitation. However, despite the low concentrations of KMD-3213 employed, we cannot rule out the possibility that blockade of a portion of the alpha 1D-AR population contributes to inhibition by KMD-3213 of responses to high PE doses. Nevertheless, the lack of effect of KMD-3213 on baseline hemodynamics and responses to low PE doses is consistent with the reported selectivity of KMD-3213 for alpha 1A-ARs at the doses used herein (1, 37).


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  		Fig. 1.   Low- and high-dose KMD-3213 (12.8 and 32 µg/kg) had no significant effect on arterial pressure or baseline renal and hindquarter resistance determined 30 min after administration to phenobarbital-anesthetized rats (top left). Control values before low- and high-dose KMD-3213 were, respectively, 104 ± 3 and 99 ± 5 mmHg and 21 ± 2 and 20 ± 2 (renal) and 38 ± 4 and 36 ± 3 (hindquarters) mmHg/kHz Doppler shift. Top right and bottom left and right: effect of KMD-3213 on increases in pressure and resistances produced by intravenous bolus doses of phenylephrine determined 30 min after KMD-3213 administration. Peak percent changes (Delta ) normalized to baseline values preceding each dose. n, Number of animals.

KMD-3213 inhibits neointimal growth. Balloon injury of the carotid artery caused the expected increases in intimal, medial, and adventitial thicknesses and areas and decreases in circumference and lumen area (Figs. 2 and 3). Thicknesses were determined to provide a measure of hypertrophy, because alterations in circumference can alter wall area independent of a change in wall thickness. Chronic intravenous infusion of low- and high-dose KMD-3213 inhibited neointimal growth. There was also a tendency, although statistically insignificant, toward inhibition of lumen loss, adventitial hypertrophy, and circumference shortening. These effects were accompanied by an absence of effect on systolic, diastolic, and mean arterial pressures and heart rate (Fig. 4; intravenous KMD-3213 administration was begun on day 0 at the time of surgery). The decline in systolic and rise in diastolic pressures after day 4 reflects the decline in degree of catheter patency with time (and complete occlusion in several animals). Direct measures obtained from new catheters on day 14 under anesthesia confirmed the absence of effect of KMD-3213. Comparison of body weight on day 0 versus day 14 among vehicle (540 ± 9 vs. 524 ± 8 g), low-dose KMD-3213 (530 ± 10 vs. 529 ± 11 g), and high-dose KMD-3213 (513 ± 11 vs. 518 ± 8 g) groups demonstrated no effect of KMD-3213.


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  		Fig. 2.   Compared with sham-injured (uninjured) right carotid, balloon injury caused expected injury response of left carotid measured 2 wk later in vehicle (Veh)-treated animals. Low- and high-dose KMD-3213 (KMD; 4 and 10 µg · kg-1 · h-1 iv) reduced neointimal area and thickness. n, Number of animals. Values were normalized to body weight. Thicknesses was calculated from areas, as a measure of hypertrophy, because of changes in circumference (see Fig. 3).



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  		Fig. 3.   Compared with sham-injured (uninjured) right carotid, balloon-injury caused expected injury response of left carotid measured 2 wk later in vehicle-treated animals. Low- and high-dose KMD-3213 (4 and 10 µg · kg-1 · h-1 iv) had no significant effect on circumference and media hypertrophy. n, Number of animals. Values were normalized to body weight. Media thicknesses was calculated from area, as a measure of hypertrophy, because of changes in circumference.



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  		Fig. 4.   Compared with vehicle, low- and high-dose KMD-3213 (4 and 10 µg · kg-1 · h-1, iv) had no effect on mean, systolic, or diastolic arterial pressures or heart rate (ANOVA). Pressures and heart rate were measured during last 5 min of a 35-min recording period done on alternate days on conscious, freely moving rats in their home cage during, beginning on day 2 after balloon injury, commencement of KMD-3213 administration on day 0. Because of progressive loss of catheter patency (indicated by narrowing of pulse pressure after day 4), values were obtained on day 14 under pentobarbital anesthesia with the use of a new catheter. n = 9 (vehicle), 10 (low-dose KMD-3213), and 8 (high-dose KMD-3213) animals at start of study (with some loss of n size due to catheter blockade after day 4). Body weights did not change over duration of experiment for the three groups (see RESULTS).


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

The major finding of this study was that chronic intravenous administration of KMD-3213 for 2 wk after balloon injury at two doses without effects on basal renal or hindquarter resistances, arterial pressures, or heart rate caused inhibition of neointimal growth in the carotid artery by 30 and 46%. Absence of hemodynamic alterations argues against the likelihood that this inhibitory effect is secondary to altered loading conditions on the injured vascular wall. (Note also that data were compared with the uninjured sham-treated right carotid artery.) These are the first in vivo findings supporting a direct trophic role for catecholamines using parenteral administration of an antagonist shown to not alter arterial pressures or regional resistances. As such, they provide strong support for other evidence, in vitro and in vivo, that suggests NE is directly trophic on SMCs and AFBs of the vascular wall, promoting proliferation, hypertrophy, migration, and collagen accumulation (8, 10, 33, 36, 38, 39). Migration of SMCs and possibly AFBs to the intima, followed by proliferation and matrix elaboration, is an essential event underlying neointimal formation after experimental balloon injury and contributes importantly to hypertrophic vascular diseases and complications after vascular surgery that result in lumen narrowing, wall hypertrophy, and fibrosis (31, 35). Besides supportive in vitro evidence (10, 36, 38, 39), we recently found by using chronic perivascular suffusion of NE and AR antagonists in rats in vivo that elevation of wall NE for 2 wk is directly trophic for the uninjured artery wall, causing inward remodeling and lumen loss (8). Moreover, NE-induced wall growth was augmented by balloon injury, such that even in the presence of basal sympathetic drive, KMD-3213-sensitive alpha 1-ARs (presumably alpha 1A), but not other alpha 1-, alpha 2- or beta -ARs (as determined by selective antagonist exposure for 2 wk), contributed significantly to experimental restenosis (8). The present study with parenteral administration extends these previous findings where dosage as well as any potential side effects of the local suffusion catheter could not be precisely known. Current findings also agree with our previous study of uninjured and balloon-injured rat aortas in organ culture where KMD-3213, but not other alpha 1-AR subtype, alpha 2- and beta -AR antagonists, blocked the effect of NE to stimulate increased protein synthesis and hypertrophy, proliferation, and reduced expression of marker proteins of the differentiated SMC phenotype in the media of uninjured rat aortas and the intima-media of balloon-injured rat aortas (39).

Conclusions from this study depend on the selectivity of KMD-3213. KMD-3213 has ~56- and 583-fold selectivity for the alpha 1A-AR over the alpha 1D- and alpha 1B-AR, respectively (1, 37). No other antagonists for alpha 1A-ARs are available with selectivities approaching that of KMD-3213, and the most selective alpha 1D-AR (BMY-7378) and alpha 1B-AR (AH11110A) antagonists available are only 180- and 20-fold selective, respectively (7, 8, 10, 13, 25, 33, 36, 38, 39). This, together with the known effect of alpha 1D-AR blockade to lower arterial pressure in rats and dogs (3, 32), precluded our use of other antagonists in the present chronic intravenous design. The acute experiment performed herein suggests that the doses of KMD-3213 we used were selective. KMD-3213 at either the low or 2.5-fold higher dose had no effect on mean arterial pressure, baseline resistance in the kidney or hindquarters, or constrictor responses evoked by "low-dose" PE. In the resting state of conscious and anesthetized rats, adrenergic contribution to baseline resistance is high in the hindquarters (predominantly skeletal muscle) (39), where it appears to be primarily dependent on alpha 1D-ARs (21, 26, 33), whereas in the kidney, it is low (9) and may be primarily alpha 1B-AR-dependent (29, 30). alpha 1A-AR-dependent constriction may be recruited in both beds during elevated sympathetic catecholamine activity (7, 13, 25, 30, 33). The hemodynamic data in the present study support these previous findings. Our current results are also consistent with the prediction from pharmacokinetic calculations that the KMD-3213 doses used herein provide functional blockade of a portion (estimated at 50-70%) of alpha 1A-ARs, with little predicted effect on alpha 1B- or alpha 1D-ARs. The absence of alterations in blood pressures and heart rates in the conscious study herein provide additional confirmation of these conclusions. It is possible that if more highly selective alpha 1A-AR antagonists were available, greater inhibitory effects on the trophic responses to balloon injury could have been demonstrated, in accordance with our findings in organ culture and with perivascular suffusion where higher KMD-3213 concentrations were used without inducing hemodynamic complications (8, 39).

We recently obtained evidence using mice with targeted disruption of catecholamine synthesis, alpha 1B- or alpha 1D-ARs, that supports the concept that NE has significant direct trophic actions on the injured vascular wall. Hypertrophic remodeling of the carotid artery induced by mechanical injury was almost abolished in mice devoid of dopamine beta -hydroxylase or alpha 1B-ARs, but was little affected in mice devoid of alpha 1D-ARs (alpha 1A-AR deficient mice have not been evaluated) (40). The absence of effect of genetic elimination of alpha 1D-ARs (40) or of alpha 1D-AR antagonist on the trophic effects of NE in our previous studies (8, 39) is noteworthy for the present study, given that the selectivity of KMD-3213 over this receptor is low (56-fold) relative to the alpha 1B-AR (583-fold). Clearly, more selective alpha 1-AR antagonists, together with genetic strategies that permit controlled stimulation and blockade of alpha 1-ARs in a vessel-specific manner, are needed to identify the full contribution of stimulation of these ARs to vascular wall growth in adaptive states, surgical complications, and vascular diseases. Interestingly, stimulation of alpha 1A-ARs may mediate hypertrophy of cardiomyocytes (13, 27) and proliferation and hypertrophy of prostate stromal cells in prostatic hypertrophy (12, 23).

In summary, intravenous administration of KMD-3213, at doses sufficient to achieve partial blockade of alpha 1A-ARs, but without systemic hemodynamic effects, significantly attenuated neointimal growth after balloon injury. The results are consistent with in vitro and in vivo studies demonstrating that alpha 1A-ARs mediate direct trophic effects of NE on SMCs of the rat aorta and carotid. Our previous organ culture studies (39) have suggested that stimulation of alpha 1B-ARs on vascular fibroblasts may also contribute to the trophic effect of NE on vascular wall growth after injury. Therefore, development of more selective antagonists permitting greater blockade of alpha 1A-ARs and/or alpha 1B-ARs, although leaving alpha 1D-ARs intact, may achieve stronger inhibition of neointimal growth, adventitial thickening, inward remodeling, and thus lumen loss, with minimal hemodynamic disturbance. Also, the present studies were conducted in animals maintained under unstressed conditions. During repeated or prolonged sympathoexcitation, adrenergic trophic contribution to intimal expansion and lumen loss may be accentuated. Future studies will be required to determine whether direct trophic effects of catecholamines contribute to other models of vascular injury and disease and to identify the cellular mechanisms underlying the growth-promoting actions of NE. Selective blockade of the trophic alpha 1-AR subtype(s) in humans may reduce diseases and surgical complications involving excessive growth of vascular wall cells.


    ACKNOWLEDGEMENTS

We thank Kirk McNaughton for histological assistance.


    FOOTNOTES

* J. C. Teeters and C. Erami contributed equally to this work.

This research was supported by National Heart, Lung, and Blood Institute Grant HL-62584.

Address for reprint requests and other correspondence: J. E. Faber, Dept. of Cell and Molecular Physiology, 474MSRB, Univ. of North Carolina, Chapel Hill, NC 27599-7545 (E-mail: jefaber{at}med.unc.edu).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

First published September 19, 2002;10.1152/ajpheart.00658.2002

Received 25 July 2002; accepted in final form 9 September 2002.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

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Am J Physiol Heart Circ Physiol 284(1):H385-H392
0363-6135/03 $5.00 Copyright © 2003 the American Physiological Society


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