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Departments of 1 Physiology and 2 Medicine, Monash University, Victoria 3800, Australia
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ABSTRACT |
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 TOP
 ABSTRACT
 INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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The hypothesis tested in this study is that diabetes has a different impact on an artery in which endothelium-dependent responses derive from both nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF) compared with responses in which NO predominates and EDHF is absent. The streptozotocin-treated rat model of diabetes was used, and the arteries were mounted on a wire myograph. In mesenteric arteries depolarized and constricted with phenylephrine, acetylcholine evoked hyperpolarization (31 ± 2 mV) and complete relaxation; these responses were attributed to EDHF and NO. In femoral arteries, acetylcholine evoked a small, NO-mediated hyperpolarization (5 ± 1 mV) and incomplete relaxation. Bradykinin evoked NO-dependent responses in mesenteric arteries. Whereas diabetes significantly impaired the EDHF-dependent hyperpolarization and relaxation in mesenteric arteries, NO-dependent responses in femoral and mesenteric arteries were preserved. 1-Ethyl-2-benzimidazolinone evoked hyperpolarization and relaxation in mesenteric arteries, and this was impaired in diabetes. In conclusion, NO-dependent responses are preserved in diabetes, whereas endothelial responses-dependent upon EDHF appear to be impaired. The putative channels responsible for mediating the EDHF response may be altered in diabetes.
hyperpolarization and relaxation; nitric oxide; endothelium-derived hyperpolarizing factor; mesenteric artery; femoral artery
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INTRODUCTION |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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THE ENDOTHELIUM is an important modulator of vascular smooth muscle tone and reactivity (8), and there is broad consensus that endothelial vasodilator dysfunction may contribute to the widespread vascular complications of diabetes mellitus (5). However, what is not clear is which vascular bed(s) or which endothelial factor(s) may be implicated in this dilator dysfunction. For example, Kiff and colleagues (11) pointed out that, whereas the blood pressure response in intact conscious diabetic rats to infusion of endothelium-dependent and endothelium-independent dilators appears normal, the responses in different individual vascular beds vary. Such differences may reflect, at least in part, differences in the contribution of the various endothelium-dependent vasodilators in different vascular beds (13, 20). For example, endothelium-dependent relaxation to acetylcholine (ACh) in the femoral artery is almost entirely dependent on nitric oxide (NO), whereas other endothelium-dependent vasodilators, especially endothelium-derived hyperpolarizing factor (EDHF), clearly play an important role in the mesenteric artery (20). Furthermore, not all agents stimulate the release of the same complement of vasodilators from endothelial cells. ACh stimulates release of NO and evokes an EDHF response in mesenteric arteries (7), whereas bradykinin stimulates predominantly NO from that vascular bed (16). These heterogeneous physiological effects of diabetes on the microcirculation may help to explain the differential impact of diabetes on the function of different vascular beds, which characterizes the clinical complications of this condition. The importance of this heterogeneity may be in directing further examination of the mechanisms underlying the specific changes in vasodilator responses in various parts of the microcirculation.
In the rat mesenteric artery a clear component of ACh-induced relaxation is resistant to blockers of NO and prostacyclin synthesis, suggesting a prominent role for EDHF in this vessel. Whereas most studies of endothelial dysfunction in diabetes have focussed on NO, Taylor and colleagues (16) reported impairment of the NO- and prostacyclin-resistant relaxation in mesenteric arteries of diabetic rats. Furthermore, the hyperpolarization attributed to EDHF in this artery is reduced in diabetes (7). These observations point to impairment of EDHF during diabetes, and we have now tested this hypothesis further by comparing endothelium-dependent responses in the mesenteric artery with those in an EDHF-independent vessel, namely, the femoral artery, which appears to depend predominantly on NO for endothelium-dependent vasodilatation. Fukao and colleagues (7) studied membrane potential in diabetic rat mesenteric arteries in the absence of depolarization and constriction, and the results suggest impairment of EDHF in this vascular bed as a consequence of diabetes. In the present study we tested this hypothesis further by comparing the effects of diabetes on endothelium-dependent vasodilatation in the femoral artery, which appears to depend solely on NO, with the responses in the mesenteric artery, in which both NO and EDHF are released from the endothelium. We recorded membrane potential simultaneously with tension in both arteries depolarized and constricted with phenylephrine to more fully elucidate the contribution by EDHF to relaxation. This is the first documentation of such simultaneous recordings in constricted arteries of diabetic rats. As an alternative approach in studying the impact of diabetes, we also compared, in an individual artery (mesenteric, see above), the hyperpolarizations and relaxations evoked by two different agonists, ACh and bradykinin, each of which releases a different suite of dilators from the endothelium.
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METHODS |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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Animals and tissues. Diabetes mellitus was induced in 8-wk-old adult male Wistar rats by injecting 60 mg/kg streptozotocin (STZ), dissolved in 0.1 M citrate buffer, into the tail vein under halothane and N2O anesthesia. Nondiabetic age-matched control rats were also anesthetized, and blood glucose levels were determined in all animals. The rats were obtained from Monash University Central Animal Services (Monash University Animal Ethics Committee approval number P540 PHY/1997/028). Each diabetic rat was housed together with its control rat, and all rats were given free access to food and water. After 8 wk the rats were anesthetized with chloroform, weighed, and killed by decapitation, and blood samples were obtained for glucose determination. The mesenteric beds and femoral arteries were removed and placed in Krebs solution containing (mM) 120.0 NaCl, 5.0 KCl, 25.0 NaHCO3, 11.0 glucose, 1.0 KH2PO4, 1.2 MgSO4, and 2.5 CaCl2, bubbled with 5% CO2-95% O2.
Distal branches of the femoral artery and tertiary branches of the mesenteric arcades (both 200- to 300-µm outside diameter) were dissected free from adipose and connective tissue, and ring segments 1-2 mm in length were separately mounted on a small vessel Mulvany-style myograph (12) for measurement of isometric tension. The arteries were continuously superfused with Krebs solution (3 ml/min) at 35°C and were stretched in increments to a tension that was equivalent to a transmural pressure of 80-90 mmHg, the mean arterial blood pressure in rats. An initial test application of the
1-adrenoceptor agonist phenylephrine (10 µM) and ACh
(10 µM) confirmed the integrity of the smooth muscle and endothelium, respectively, and the preparations were then allowed to recover for 30 min.
Contraction and endothelium-independent relaxation. Cumulative concentration-contraction curves to phenylephrine were performed to assess the ability of the muscle to contract. The ability of the smooth muscle to relax independently of the endothelium was tested using cumulative concentration-relaxation curves to sodium nitroprusside (SNP).
Endothelium-dependent relaxation.
The arteries were submaximally constricted with phenylephrine.
Increasing concentrations of ACh (1 nM-10 µM) were applied for 2 min each with a washout between each dose sufficient to allow the
tension to return to its prerelaxation level as described previously
(15). After this initial concentration-relaxation curve
was completed, the phenylephrine was washed out, and the artery was
incubated with the inhibitor of NO synthase
N
-nitro-L-arginine methyl ester
(L-NAME), 50 or 100 µM, for 30 min. In the continued
presence of phenylephrine and L-NAME, the cyclooxygenase
inhibitor indomethacin (1 µM) was added to the superfusate and, after
30 min, a third concentration-relaxation curve to ACh was performed.
Recording of membrane potential.
In a separate series of experiments the protocol described above was
repeated but, in addition, the membrane potential of the smooth muscle
cells was recorded simultaneously with tension using conventional glass
intracellular microelectrodes filled with 1 M KCl with resistances of
60-100 M
, as described previously (14, 15). This
enabled quantification of the hyperpolarization associated with
ACh-evoked relaxation in both mesenteric and femoral arteries.
Initial control experiments. From the concentration-response curves to phenylephrine, the concentrations of that drug that yielded similar levels of submaximal contraction were determined. Phenylephrine (3 µM) produced 79 ± 3% and 81 ± 4% contraction in femoral arteries from control and diabetic rats, respectively, and 10 µM induced 78 ± 4% and 78 ± 3% contraction in mesenteric arteries from control and diabetic rats, respectively (n = 10 rats in all 4 groups).
Because each tissue was subjected to three concentration-response curves, control, plus L-NAME, and L-NAME plus indomethacin, a series of time-control experiments without L-NAME or indomethacin was run. Three concentration-relaxation curves, with 30-min intervening, were indistinguishable. The concentration of L-NAME required to achieve a reproducible rightward shift in the concentration-relaxation curve in response to ACh was verified in mesenteric arteries from five control rats. The pD2 for ACh-evoked relaxation was 7.16 ± 0.08 and the maximum response (Vmax) was 11 ± 5% of the phenylephrine-induced contraction (n = 7) in the presence of 50 µM L-NAME. Increasing the concentration of L-NAME to 100 µM shifted the pD2 to 7.02 ± 0.14 (P > 0.05) and Vmax to 20 ± 7% (P = 0.36), which were not significantly different compared with the lower concentration of L-NAME. Increasing the concentration of L-NAME to 1 mM had no further effect. Finally, in most previous studies, ACh has been applied cumulatively when stimulating endothelium-dependent relaxation. When recording membrane potential simultaneously with tension, we have routinely applied ACh discretely, each application lasting 1 or 2 min with a return of the tension to its prerelaxation level between each application (15). Using mesenteric arteries obtained from five control rats, we constructed concentration-relaxation curves to ACh applied both cumulatively and discretely to each tissue. Both curves were identical using the two methods.Histology. Segments of femoral and mesenteric arteries from diabetic and control rats were dissected out, fixed in 10% phosphate-buffered formalin, mounted in paraffin blocks, and cut into 10-µm slices for staining with Masson's trichrome. This stain distinguishes connective tissue from smooth muscle cells and therefore enabled verification of the muscular status of the arteries that were studied.
Drugs. Drugs used in this study were the following: STZ, phenylephrine hydrochloride, ACh hydrochloride, L-NAME, indomethacin, bradykinin acetate, apamin, and superoxide dismutase (S-2515 from bovine erythrocytes) (Sigma; St. Louis, MO); charybdotoxin (Auspep, Melbourne, Australia), SNP (Ajax Chemicals; Auburn, New South Wales, Australia); and 1-ethyl-2-benzimidazolinone (EBIO, Tocris, Bristol, UK). Phenylephrine, ACh, L-NAME, and bradykinin were prepared as stock solutions in distilled water, indomethacin in Na2CO3 (0.1 M), and EBIO in dimethyl sulfoxide.
Statistical analysis.
Relaxations were expressed as a percentage of the contraction induced
by phenylephrine, and hyperpolarization was expressed as the change in
membrane potential (mV). Concentration-response curves were calculated,
and a sigmoid curve was fitted to data for each artery using the
least-squares method (Prism, Graphpad Software). The concentration of
agonist that was effective in producing a response that was 50% of the
maximum response (EC50), the maximum response induced by
ACh (Vmax), and the slope were determined for
each curve. For each group of data the mean values for
EC50, Vmax, and slope means ± SE for each concentration of agonist were calculated, and a curve
representative of each group was produced. pD2 (
log
EC50) values have been quoted throughout when comparing
responses of arteries to ACh, bradykinin, phenylephrine, and SNP.
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RESULTS |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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Blood glucose levels and animal weights. At the time of the experiment, all STZ-treated rats exhibited hyperglycemia with blood glucose concentrations (31.0 ± 1.5 mmol/l, n = 10) significantly higher than those of the age-matched nondiabetic control rats (5.8 ± 0.5 mmol/l, n = 9, P < 0.0001). The weights of the rats at the time of injection were similar (293 ± 9 g, n = 10, for the diabetic rats; 281 ± 9 g for the controls, n = 9, P = 0.37), but the final weight of the diabetic rats (309 ± 14 g, n = 10) was significantly lower than that of the control rats (501 ± 23 g, n = 9).
ACh-induced relaxation in mesenteric versus femoral
arteries of control rats.
In nondiabetic control rats, endothelium-dependent relaxation of the
femoral arteries required higher concentrations of ACh (pD2, 6.52 ± 0.07, n = 5) than the
mesenteric arteries (pD2, 7.62 ± 0.04, n = 9, P < 0.001). Whereas the highest
concentrations of ACh completely relaxed the mesenteric artery (only
1 ± 3% of the phenylephrine-induced contraction remained,
n = 9), the maximum tension remaining in the femoral
artery was 43 ± 9% (n = 5, P =
0.01) (Fig. 1).
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Effect of L-NAME and indomethacin on
ACh-induced vasodilatation in arteries of control rats.
L-NAME (50 µM) abolished endothelium-dependent relaxation
to ACh in the femoral artery (Fig. 2).
This contrasted with observations in the mesenteric artery in which
almost complete relaxation persisted in the presence of
L-NAME, although there was a significant rightward shift in
the pD2 (P < 0.0001) (Table
1, Fig.
3B). The addition of
indomethacin, in the continued presence of L-NAME, had no
further impact on endothelium-dependent relaxation to ACh in the
mesenteric artery (Table 1, Fig. 3B).
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Diabetes and ACh-induced vasodilatation. In femoral arteries, there was no difference between the concentration-relaxation curves to ACh in tissues from diabetic rats compared with nondiabetic control tissues (Table 1, Fig. 2). Likewise, L-NAME abolished the relaxation response to ACh in tissues from diabetic rats as it had in controls (Fig. 2).
In mesenteric arteries in control solution, the concentration-relaxation curves to ACh were shifted to the right in arteries from diabetic rats compared with those from nondiabetic controls, but the Vmax was not different and almost complete relaxation of the phenylephrine-induced contraction was achieved in both groups (Table 1, Fig. 3A). In the presence of L-NAME, the pD2 for ACh-induced relaxation in mesenteric arteries from diabetic rats was shifted significantly to the right (Table 1, Fig. 3B). The shift in pD2 in the diabetic arteries by L-NAME was 0.50 ± 0.19 (n = 10), which was not different from the shift of 0.42 ± 0.08 (n = 9) in controls (P = 0.10, Table 1). However, the reduction in Vmax by L-NAME was significantly greater in the mesenteric arteries from the diabetic rats compared with controls (Table 1, Fig. 3B). L-NAME reduced Vmax by only 6 ± 5% in controls (n = 9) compared with 35 ± 10% in diabetics (n = 10, P = 0.02). Incubation in the free radical scavenger superoxide dismutase (150 U/ml for 20 min before and during testing) failed to improve the concentration-relaxation curves in response to ACh (n = 5) in mesenteric arteries obtained from either control or diabetic rats (data not shown). Indomethacin was without further effect on the concentration-relaxation curves (pD2 or Vmax) for ACh in mesenteric arteries from diabetic rats (Table 1, Fig. 3B). The maximum relaxation evoked by ACh in mesenteric arteries in the presence of L-NAME plus indomethacin was abolished by 5 min exposure to charybdotoxin (30 nM) plus apamin (0.25 µM) (n = 3). The effect of these blockers was reversed 30 min after they had been washed out.Hyperpolarization evoked by ACh in mesenteric and
femoral arteries.
The resting membrane potentials of the smooth muscle cells were not
different in mesenteric arteries of diabetic rats (58 ± 2 mV,
n = 6) compared with controls (59 ± 2 mV,
n = 6). When the arteries were depolarized and
constricted with phenylephrine, the membrane potentials were also
similar (diabetic, 34 ± 1 mV, n = 6; control,
35 ± 1 mV, n = 5).
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Bradykinin-induced responses and the effect of diabetes.
In mesenteric arteries, there was no difference between the
concentration-relaxation curves to bradykinin in tissues from diabetic
rats versus controls (Table 1, Fig. 6).
L-NAME markedly reduced the bradykinin-evoked relaxation in
arteries from both diabetic rats and controls (Table 1, Fig. 6).
Addition of indomethacin, in the presence of L-NAME,
abolished all bradykinin-induced relaxation (Table 1, Fig. 6).
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Effects of EBIO.
A significant proportion of the hyperpolarization evoked in the
mesenteric artery by ACh in this study persisted in the presence of
L-NAME plus indomethacin and hence was attributed to EDHF. Because the EDHF response is blocked by the combination of
charybdotoxin and apamin in a variety of vascular smooth muscles
(4, 6, 19), it has been ascribed to the activation of
intermediate- and small-conductance, Ca2+-activated
K+ channels. The intermediate-conductance channels can be
activated by EBIO (10). In the present study, EBIO evoked
concentration-dependent hyperpolarization and relaxation in tissues
obtained from control rats, whereas the responses in tissues from
diabetic rats were absent or significantly reduced (Fig.
7).
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Endothelium-independent constriction and relaxation of arterial smooth muscle in diabetic and control rats. The ability of phenylephrine to evoke vasoconstriction was not different in the mesenteric arteries from diabetic (pD2, 5.60 ± 0.03, n = 6) compared with control rats (pD2, 5.52 ± 0.02, n = 8, P = 0.08). Thus the ability of the smooth muscle to contract was unaffected by diabetes.
Likewise, the ability of the smooth muscle to relax was preserved in arteries obtained from diabetic rats. In mesenteric arteries, the pD2 for SNP-induced relaxation was 7.45 ± 0.07 (n = 5) for control rats and 7.57 ± 0.07 (n = 5) in tissues from diabetic rats (P = 0.26). Vmax values for these responses were 3 ± 4% and 2 ± 3% of the phenylephrine-induced contraction, respectively (P = 0.85). In femoral arteries, the relaxations evoked by 10 µM SNP were 64 ± 9% (n = 5) and 79 ± 8% (n = 4) in control and diabetic rats, respectively (P = 0.26).Histology. The media of both femoral and mesenteric arterial walls stained red with Masson's trichrome indicating their status as muscular arteries. In the control rats the muscle layer was 6.4 ± 0.2 smooth muscle cells thick in the femoral artery (n = 8) and 4.3 ± 0.3 cells thick in the mesenteric artery (n = 8). In the diabetic rats the average number of muscle cells in the wall was 6.3 ± 0.3 in the femoral artery (n = 4) and 4.0 ± 0.3 in the mesenteric artery (n = 7), not different from the controls. Whereas in the dissecting dish in the absence of pressure, the outside diameter of all the arteries studied was 200-300 µm.
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DISCUSSION |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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This study demonstrates that the impact of STZ-induced diabetes on the vasodilator function and control of the resistance circulation in rats is heterogeneous. The importance of this heterogeneity is that it appears to reflect an impairment of the functional response to EDHF rather than NO. Evidence to support this conclusion includes the finding that L-NAME has a greater impact on ACh-evoked maximum relaxation and hyperpolarization in mesenteric arteries from diabetic rats than from controls, suggesting that NO bioavailability, at least in vitro, is not reduced in the arteries of diabetic rats. Furthermore, definitive evidence provided by simultaneous recordings of membrane potential and tension confirmed that EDHF-dependent hyperpolarization and the associated relaxation is markedly reduced in mesenteric arteries from diabetic rats compared with controls. Finally, relaxation that relies predominantly on the release of NO from the endothelium, i.e., ACh-induced relaxation in the femoral artery and bradykinin-induced relaxation in the mesenteric artery, is resistant to the effects of diabetes. This implicates EDHF rather than NO in the endothelium-dependent vasodilator dysfunction in this model of diabetes.
Most studies examining the impairment of endothelial vasodilator function associated with diabetes have focused on the role of NO, and conflicting conclusions have been reached (see Ref. 5). The greater reduction of both maximum relaxation and maximum hyperpolarization by L-NAME in mesenteric arteries of diabetic rats suggests a greater dependence on NO in diabetes. Thus our results support those that suggest NO production may be unchanged or even increased in mesenteric arteries of diabetic rats (see Refs. 5 and 9). An increase in the burden of destructive reactive oxygen species in diabetes (2, 17) in vivo would reduce the bioavailability of NO, which could stimulate an increase in NO production (see Ref. 5).
The prominence of L-NAME-resistant relaxation to ACh in the mesenteric artery provides strong evidence that endothelium-dependent vasodilators in addition to NO are important in these vessels (7, 18, 20). Our electrophysiological recordings confirm that EDHF plays an important role in the ACh-evoked, endothelium-dependent relaxation of mesenteric arteries, and that this pathway is compromised following 8 wk of diabetes. This conclusion supports the observations of Fukao and colleagues (7) who recorded membrane potential in rat mesenteric arteries pinned to the bottom of a recording bath and not exposed to constrictor. They showed that the amplitude and duration of the hyperpolarization produced by acetylcholine was significantly decreased in arteries of diabetic rats. Blockade of NO synthesis did not affect the hyperpolarizing response to ACh in either diabetic animals or controls in that study and, furthermore, in vitro application of superoxide dismutase, a scavenger of superoxide anions, was without effect on the ACh-induced hyperpolarization. In our study L-NAME reduced the hyperpolarization in the diabetic arteries. The 31-mV hyperpolarization attributed to EDHF dominated the membrane potential response in control arteries and probably masked the smaller NO-dependent hyperpolarization. In the diabetic arteries the EDHF response was markedly reduced, revealing the NO-dependent hyperpolarization, and this hyperpolarization was reduced by L-NAME. NO-dependent hyperpolarization is sensitive to the degree of stretch on the artery (14) and thus more likely to be detected in arteries mounted on a myograph than in arteries pinned to the base of a recording chamber. Nonetheless, our results confirm and extend Fukao et al.'s initial observations (7), lending support to the notion that the functional EDHF response is decreased in mesenteric arteries during diabetes in rats. Simultaneous measurement of membrane potential and tension in our study enabled us to unequivocally demonstrate that impaired hyperpolarization underpinned the reduced ACh-induced relaxation in these arteries.
We have also tested the hypothesis further by recording membrane potential simultaneously with tension in the femoral artery, in which endothelium-dependent vasodilation appears to depend predominantly, if not solely, on NO. In this vessel the maximum hyperpolarization induced by ACh was only 5 mV in amplitude, typical of NO rather than EDHF (14), and persisted (4 mV) in diabetes. The hyperpolarization and relaxation in the femoral artery were completely blocked by L-NAME.
The incompleteness of the endothelium-dependent relaxation in the femoral artery (43%) is unlikely to reflect additional destruction of NO in the thicker arterial wall, because the relaxation achieved by SNP was also less than maximal (36%). Interestingly, the maximal relaxation evoked by bradykinin in the mesenteric artery was equivalent to that evoked by ACh in the femoral artery; both were submaximal and dependent on NO.
Our observation in femoral arteries that ACh-induced hyperpolarization and relaxation is dependent on NO and entirely resistant to the effects of diabetes supports our conclusion that reduced EDHF rather than NO is responsible for the endothelial vasodilator dysfunction in diabetes. Only one other in vitro study has investigated the impact of diabetes on endothelium-dependent relaxation in a hindlimb vessel of the rat. Taylor and colleagues (16) examined endothelium-dependent responses to ACh in mesenteric and popliteal arteries of rats with diabetes of short duration (3 wk), and they demonstrated impairment of relaxation in both arteries, but blockers were not used on the diabetic arteries to verify the nature of the factor involved. Taylor and colleagues (16) also found no impact of diabetes on endothelium-dependent relaxation to bradykinin in either mesenteric or popliteal arteries, and our observation supports that finding. Furthermore, we also demonstrated that bradykinin-evoked relaxation in arteries from both control and diabetic rats were abolished by L-NAME. We interpret these observations in terms of preservation of the NO-dependent relaxation in STZ-induced diabetes (i.e., ACh-evoked relaxation in femoral arteries and bradykinin-evoked relaxation in mesenteric arteries are resistant to diabetes). This supports our hypothesis that a NO-independent vasodilator such as EDHF is vulnerable to the effects of diabetes.
The hyperpolarization attributed to EDHF is abolished by charybdotoxin and apamin in a variety of arteries (4, 6, 19), suggesting the involvement of intermediate- and small-conductance, calcium-activated potassium channels (1, 3). We have confirmed that the EDHF-attributed relaxation in the mesenteric artery is also blocked by charybdotoxin and apamin. An activator of the intermediate-conductance channels EBIO (10) induced hyperpolarization and relaxation in mesenteric arteries of control rats but was ineffective in tissues from diabetic rats. This could reflect a reduction in ion channel density or function in endothelial cells in diabetes. Whether this is due to a direct toxic effect of hyperglycemia or reflects a more complex consequence of diabetes is uncertain. In any case, our results suggest that, whereas attempts to increase NO bioavailability may be of limited value in improving endothelial vasodilator function in diabetes, at least in the mesenteric and femoral arteries of STZ-induced diabetic rats, further examination of the above-mentioned endothelial cell intermediate- and small-conductance, calcium-activated potassium channels may be important in our understanding of the heterogeneity of the impact of diabetes on the microcirculation and the cellular mechanisms underlying the vasodilator impairment at different sites.
In conclusion, we have shown that there is a selective impairment of endothelium-dependent relaxation to ACh in mesenteric, but not in femoral, arteries rather than a generalized impairment across all resistance circulations, and this impairment is attributable to reduced EDHF-dependent rather than NO-dependent responses. Impairment in diabetes is most pronounced in situations in which EDHF contributes substantially to endothelium-dependent relaxation (i.e., ACh-induced relaxation in mesenteric arteries), whereas NO-dependent relaxation (i.e., ACh-induced relaxation in femoral arteries and bradykinin-induced relaxation in mesenteric arteries) is resistant to the effects of diabetes. In addition, an individual artery in which endothelium-dependent relaxation induced by one agent is impaired during diabetes may be resistant to diabetes with respect to endothelium-dependent vasorelaxation induced by another agent. This study has thus improved our understanding of the relative contributions and deficiencies of the three endothelium-dependent vasodilators in the endothelial vasodilator dysfunction associated with diabetes, at least that which is induced in rats by STZ. Our use of intracellular microelectrodes to measure hyperpolarization and our comparison of two different resistance arterial beds within the same animal has confirmed that the hyperpolarization, the defining characteristic of EDHF, is markedly reduced in this form of diabetes and that this hyperpolarization underpins relaxation. Such understanding is critical to the development of clinically useful intervention strategies to limit the vascular complications of diabetes.
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ACKNOWLEDGEMENTS |
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The authors thank Joanne Waring (Department of Physiology, Monash University) for assistance with STZ injections of rats.
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FOOTNOTES |
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This work was supported by the National Health and Medical Research Council of Australia and the National Heart Foundation of Australia.
Address for reprint requests and other correspondence: H. C. Parkington, Dept. of Physiology, PO Box 13F, Monash Univ., Victoria 3800, Australia (E-mail: h.parkington{at}med.monash.edu.au).
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.
Received 17 November 2000; accepted in final form 8 March 2001.
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REFERENCES |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
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