Pial Arteriolar Dilation Research Articles

Beta-Adrenoceptor and nNOS-derived NO interactions modulate hypoglycemic pial arteriolar dilation in rats.

We examined the relative contributions from nitric oxide (NO) and catecholaminergic pathways in promoting cerebral arteriolar dilation during hypoglycemia (plasma glucose congruent with 1.4 mM). To that end, we monitored the effects of beta-adrenoceptor (beta-AR) blockade with propranolol (Pro, 1.5 mg/kg iv), neuronal nitric oxide synthase (nNOS) inhibition with 7-nitroindazole (7-NI, 40 mg/kg ip) or ARR-17477 (300 microM, via topical application), or combined intravenous Pro + 7-NI or ARR-17477 on pial arteriolar diameter changes in anesthetized rats subjected to insulin-induced hypoglycemia. Additional experiments, employing topically applied TTX (1 microM), addressed the possibility that the pial arteriolar response to hypoglycemia required neuronal transmission. Separately, Pro and 7-NI elicited modest but statistically insignificant 10-20% reductions in the normal ~40% increase in arteriolar diameter accompanying hypoglycemia. However, combined Pro-7-NI was accompanied by a >80% reduction in the hypoglycemia-induced dilation. On the other hand, the combination of intravenous Pro and topical ARR-17477 did not affect the hypoglycemia response. In the presence of TTX, the pial arteriolar response to hypoglycemia was lost completely. These results suggest that 1) beta-ARs and nNOS-derived NO interact in contributing to hypoglycemia-induced pial arteriolar dilation; 2) the interaction does not occur in the vicinity of the arteriole; and 3) the vasodilating signal is transmitted via a neuronal pathway.

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Nitric oxide donor-induced increase in permeability of the blood–brain barrier

The goal of the present study was to determine the effect of nitric oxide (NO) donors on the permeability of the blood–brain barrier in vivo. We examined the pial microcirculation in rats using intravital fluorescence microscopy. Permeability of the blood–brain barrier was quantitated by calculating the clearance of fluorescent-labeled dextran ( M w=10 000 Da; FITC–dextran-10K) during suffusion with vehicle, S-nitroso- N-acetylpenicillamine (SNAP; 100 μM) and 3-morpholinosydnonimin (SIN-1; 100 μM). In addition, we examined changes in arteriolar diameter during suffusion with vehicle, SNAP and SIN-1. During suffusion with vehicle, clearance of FITC–dextran-10K from pial vessels and diameter of pial arterioles remained relatively constant during the experimental period. In contrast, suffusion with SNAP or SIN-1 markedly increased clearance of FITC–dextran-10K from the cerebral microcirculation and produced a rapid, sustained dilatation of pial arterioles. Thus, NO donors increase the permeability of the blood–brain barrier and produce pronounced dilatation of cerebral arterioles. In light of evidence suggesting that NO donors may produce their effect by the simultaneous release of NO and superoxide anion to form peroxynitrite, we elected to examine the role of superoxide anion in increases in permeability of the blood–brain barrier in response to SNAP and SIN-1. We found that suffusion with tiron (1 mM) did not alter basal permeability of the blood–brain barrier, but significantly inhibited increases in permeability of the blood–brain barrier in response to SNAP and SIN-1. In addition, tiron did not alter baseline diameter of cerebral arterioles, or SNAP- and SIN-1-induced cerebrovasodilatation. The findings of the present study suggest that NO donors produce an increase in permeability of the blood–brain barrier which appears to be related to the presence of NO and superoxide anion, to presumably form peroxynitrite. We suggest that increases in NO formation observed during brain trauma may contribute to disruption of the blood–brain barrier.

L-arginine partially restores the diminished CO2 reactivity after mild controlled cortical impact injury in the adult rat.

Using an open cranial window technique, the authors investigated the mechanisms associated with the suppressed CO2 reactivity after mild controlled cortical impact (CCI) injury in rats. The dilation of arterioles (n = 7) to hypercapnia before injury was 38 +/- 12%, which was significantly reduced both at 1 hour (23 +/- 15% dilation) and at 2 hours after injury (11 +/- 19% dilation). In the presence of L-arginine (10 mmol/L topical suffusion, 300 mg/kg intravenous infusion), the dilation of pial arterioles (n = 6) to hypercapnia was partially restored to 30 +/- 6% at 2 hours after injury. In the presence of the nitric oxide (NO) donor, S-nitroso-N-acetylpenicillamine (SNAP) (10(-8) mol/L topical suffusion), the dilation of pial arterioles (n = 5) to hypercapnia remained diminished (5 +/- 7%) at 2 hours after injury. The dilation of pial arterioles (n = 4) to hypercapnia also remained suppressed (5 +/- 6%) with topical suffusion of the free radical scavengers, polyethylene glycol-superoxide dismutase (60 units/mL) and polyethylene glycol-catalase (40 units/mL). The authors have shown that L-arginine at least partially restores the diminished response to hypercapnia after mild CCI injury. Furthermore, these data suggest that the beneficial effects of L-arginine are mediated by a combination of providing substrate for NO synthase and scavenging free radicals.

Hyposerotonin-induced nitric oxide supersensitivity in the cerebral microcirculation.

To investigate the relationship between hyposerotonin and cranial microvascular responses to nitric oxide (NO). Although the mechanism underlying NO supersensitivity in migraine is still unclear, an alteration of the serotonin system is a possible explanation. Wistar rats were divided into control and hyposerotonin groups. Serotonin was depleted by intraperitoneal injection with 300 mg/kg of para-chlorophenylalanine (PCPA), a tryptophan hydroxylase inhibitor. Three days after PCPA pretreatment, the animals were prepared for assessment of their NO-induced vasomotor response using glyceryl trinitrate (GTN: 8 to 10 mg/kg, intravenously) as an NO donor. Pial circulation was visualized by the intravital fluorescein videomicroscopic technique. Images of vessels at 0, 5, 15, 30, and 60 minutes post GTN infusion were digitized and measured. At the end of monitoring, the rat brains were removed for ultrastructural study of the brain microvessels. Infusion of GTN produced dose-dependent pial arteriolar dilatation. This vasodilator effect was significantly increased in the PCPA-treated groups, especially at 30 and 60 minutes. The percentage change from baseline diameter at 30 minutes after the 8-mg/kg GTN infusion was 42.6 +/- 3.1 for the hyposerotonin group and 16.8 +/- 2.9 for the control group (P<.001). Electron microscopic study revealed that exposure to the NO donor produced considerable changes in cerebral microvessels, characterized by focal ballooning of endothelial cells, increased microvillous formation, and increased endothelial pinocytosis. These anatomical changes were significantly more prominent in the hyposerotonin group. A hyposerotoninergic condition can facilitate the NO-induced physiological and pathological responses in meningeal and cerebral microvessels and, therefore, is a possible explanation for the supersensitivity to NO observed in patients with migraine.

Preservation of cerebrovascular tone and reactivity by sodium channel inhibition in experimental prolonged asphyxia in piglets.

Sodium channels using cAMP as a second messenger play a role in the regulation of cerebral circulation and metabolism. Cerebrospinal fluid (CSF) cAMP levels have been shown to correlate with the degree and duration of hypoxic injury and outcome and to be an indicator of cerebral vascular reactivity. We hypothesize that sodium channel inhibition either before or at termination of experimental asphyxia will attenuate cerebrovascular alterations and maintain CSF cAMP levels. Three groups of piglets with closed cranial windows were studied: asphyxia or group 1 (n = 5) and two treatment groups. Pigs were treated with 50 mg/kg of sodium channel blocker before asphyxia (group 2, n = 6) and after the termination of asphyxia and start of reventilation (group 3, n = 6). Asphyxia was sustained over 60 min by ventilating piglets with 10% O2 gas mixture and decreasing minute ventilation followed by 60 min of reventilation with room air. Every 10 min, pial arterial diameters were measured, and CSF samples were collected for cAMP determination. Vascular reactivity to topically applied isoproterenol (10(-4) M) was evaluated 60 min after recovery. During asphyxia, cAMP levels in group 2 peaked and declined at a later time with mean values remaining significantly higher than those of groups 1 and 3. During reventilation, CSF cAMP concentrations were highest in group 3 and lowest in group 1. Pial arteriolar dilation occurred during asphyxia in all three groups but to a lesser degree in the pretreated group compared with groups 1 and 3. Pial arteriolar reactivity to isoproterenol postasphyxia was preserved in both groups 2 and 3. In summary, in newborn pigs, pretreatment with sodium channel blocker resulted in higher CSF cAMP levels and a lesser degree of pial arteriolar dilation during prolonged asphyxia. Pretreatment or treatment at reventilation restored vascular tone and reactivity.

Nitric-Oxide-Dependent Pial Arteriolar Dilation in the Female Rat: Effects of Chronic Estrogen Depletion and Repletion

In this study, we compared endothelial nitric oxide synthase (eNOS)-mediated cerebral vasodilating responses in intact female rats, chronically ovariectomized (OVX) rats, and OVX rats treated for 2 weeks with 17β-estradiol (E2). Under anesthesia, using intravital microscopy and a closed cranial window system, pial arteriolar diameter changes were monitored during sequential cortical suffusions of an eNOS-dependent dilator [acetylcholine (ACh)] and a direct NO donor [S-nitrosoacetylpenicillamine (SNAP)]. In separate rats from the same groups, we compared eNOS and caveolin-1 (CAV-1) protein abundance in pial arterioles (via immunofluorescence analyses). In untreated and low-dose E2-treated (1.0 μg · kg−1 · day−1) OVX rats, ACh-induced vasodilations were virtually absent. High-dose E2 treatment (100 μg · kg−1 · day−1) restored ACh-induced pial arteriolar dilations to levels seen in intact females. The vasodilations elicited by SNAP and ADO were unaffected by chronic estrogen changes, indicating no direct estrogen influence on vascular smooth muscle (VSM) reactivity. Pial arteriolar eNOS protein abundance was diminished by ovariectomy and restored by high-dose E2 treatment. Pial arteriolar CAV-1 expression was higher in OVX versus intact and E2-treated OVX females. These results suggest that long-term changes in estrogen directly influence brain eNOS functional activity. The estrogen-related changes in eNOS-dependent vasodilating function appear to be related, in part, to a capacity for E2 to increase eNOS protein expression and, in part, to an E2-associated diminution in endothelial CAV-1 expression.

Mitochondrial potassium channel opener diazoxide preserves neuronal-vascular function after cerebral ischemia in newborn pigs.

N-Methyl-D-aspartate (NMDA) elicits neuronally mediated cerebral arteriolar vasodilation that is reduced by ischemia/reperfusion (I/R). This sequence has been preserved by pretreatment with the ATP-sensitive potassium (K(ATP)) channel opener aprikalim, although the mechanism was unclear. In the heart, mitochondrial K(ATP) channels (mitoK(ATP)) are involved in the ischemic preconditioning-like effect of K(+) channel openers. We determined whether the selective mitoK(ATP) channel opener diazoxide preserves the vascular dilation to NMDA after I/R. Pial arteriolar diameters were determined with the use of closed cranial window/intravital microscopy in anesthetized piglets. Vascular responses to NMDA were assessed before and 1 hour after 10 minutes of global cerebral ischemia induced by raising intracranial pressure. Subgroups received 1 of the following pretreatments before I/R: vehicle; 1 to 10 micromol/L diazoxide; and coapplication of 100 micromol/L 5-hydroxydecanoic acid (5-HD), a K(ATP) antagonist with diazoxide. NMDA-induced dose-dependent pial arteriolar dilation was not affected by diazoxide treatment only but was severely attenuated by I/R. In contrast, diazoxide dose-dependently preserved the NMDA vascular response after I/R; at 10 micromol/L, diazoxide arteriolar responses were unaltered by I/R. The effect of diazoxide was antagonized by coapplication of 5-HD with diazoxide. Percent preservation of 100 micromol/L NMDA-induced vasodilation after I/R was 53+/-19% (mean+/-SEM, n=8) in vehicle-treated controls versus 55+/-10%, 85+/-5%, and 99+/-15% in animals pretreated with 1, 5, and 10 micromol/L diazoxide (n=8, n=8, and n=12, respectively) and 60+/-15% in the group treated with 5-HD+diazoxide (n=5). The mitoK(ATP) channel opener diazoxide in vivo preserves neuronal function after I/R, shown by pial arteriolar responses to NMDA, in a dose-dependent manner. Thus, activation of mitoK(ATP) channels may play a role in mediating the protective effect of other K(+) channel openers.

Miconazole represses CO2-induced pial arteriolar dilation only under selected circumstances

Previous experimental findings have led to the suggestion that guanosine 3',5'-cyclic monophosphate (cGMP) plays a permissive role in hypercapnic cerebral vasodilation. However, we recently reported that the technique used to reveal a permissive role for cGMP [cGMP repletion in the presence of nitric oxide synthase (NOS) inhibition] created a situation where CO(2) reactivity was normalized but where different mechanisms (i.e., K(+) channels) participated in the response. In the present study, we examined whether that nascent K(+)-channel dependence is related in any way to an increase in the influence of the miconazole-inhibitable cytochrome P-450 epoxygenase pathway. Using intravital microscopy and a closed cranial window system in adult rats, we measured pial arteriolar diameters during normo- and hypercapnia, first in the absence and then in the presence of a neuronal NOS (nNOS) inhibitor [7-nitroindazole (7-NI)]. This was followed by suffusion of a cGMP analog and then cGMP plus miconazole. Separate groups of rats were used to evaluate whether miconazole either alone or in the presence of 8-bromoguanosine 3', 5'-cyclic monophosphate (8-BrcGMP) or its vehicle (0.1% ethanol) had any effect on CO(2) reactivity and whether miconazole affected K(+)-channel opener-induced dilations. Hypercapnic (arterial PCO(2), congruent with65 mmHg) pial arteriolar dilations, as expected, were reduced by 70-80% with 7-NI and restored with cGMP repletion. CO(2) reactivity was again attenuated after miconazole introduction. Miconazole, with and without 8-BrcGMP, and its vehicle had no influence on pial arteriolar CO(2) reactivity in the absence of nNOS inhibition combined with cGMP repletion. Miconazole alone also did not affect vasodilatory responses to K(+)-channel openers. Thus present results suggest that the nascent K(+)-channel dependence of the hypercapnic response found in our earlier study may be related to increased epoxygenase activity. The specific reasons why the pial arteriolar CO(2) reactivity gains a K(+)-channel and epoxygenase dependence only under conditions of nNOS inhibition and cGMP restoration remain to be identified. These findings again call into question the interpretations applied to data collected in studies evaluating potential permissive actions of cGMP or NO.

Cycloheximide rapidly inhibits cortical COX activity and COX-dependent pial arteriolar dilation in piglets.

We have previously shown that cycloheximide (CHX) preserved neuronal function after global cerebral ischemia in piglets, in a manner similar to indomethacin. To elucidate the mechanism of this protection, we tested the hypothesis that CHX would inhibit cyclooxygenase (COX) activity in the piglet cerebral cortex and vasculature. Pial arteriolar responses to hypercapnia, arterial hypotension, and sodium nitroprusside (SNP) were determined before and 20 min after treatment with CHX (0.3-1 mg/kg iv) using a closed cranial window and intravital microscopy. We also determined baseline and arachidonic acid (AA)-stimulated cortical PGF(2alpha) and 6-keto-PGF(1alpha) production before and 20-60 min after CHX (1 mg/kg iv) treatment, using ELISA kits. CHX did not affect baseline diameters (approximately 100 microm) but significantly decreased arteriolar dilation to COX-dependent stimuli, such as hypercapnia and hypotension, but not to COX-independent SNP. In the 1 mg/kg CHX-treated group, increases in vascular diameters were reduced from 22 +/- 2 to 10 +/- 2%, from 49 +/- 5 to 31 +/- 3% (means +/- SE, 5 and 10% CO2, respectively, n = 8), from 12 +/- 3 to 3 +/- 1%, and from 26 +/- 5 to 6 +/- 2% ( approximately 25 and 40% decreases in blood pressure, respectively, n = 6). CHX also inhibited conversion of exogenous AA to both PGF(2alpha) and 6-keto-PGF(1alpha); for example, 20 min after CHX treatment 10 microg/ml AA-stimulated PGF(2alpha) concentrations in the artificial cerebrospinal fluid decreased from 14.28 +/- 3.04 to 5.90 +/- 1.26 ng/ml (n = 9). Thus CHX rapidly decreases COX activity in the piglet cerebral cortex. This result may explain in part the preservation of neuronal function of CHX in cerebral ischemia.

Nociceptin/orphanin FQ dilates pial arteries by K ATP and K ca channel activation

Nociceptin/orphanin FQ (NOC/oFQ) is a recently discovered endogenous ligand for the opioid like receptor, ORL-1. In the piglet, cGMP activates the ATP sensitive (K ATP) while cAMP activates both the K ATP and the calcium sensitive (K ca) K + channel to elicit vasodilation. The present study was designed to characterize the role of cGMP, cAMP, K ATP, and K ca channel activation in NOC/oFQ-induced pial artery dilation in newborn pigs equipped with a closed cranial window. NOC/oFQ (10 −8, 10 −6 M) induced pial arteriole dilation was decreased by the protein kinase A inhibitor Rp 8-Br cAMPs (16±1 and 30±1 vs. 5±1 and 10±1%). NOC/oFQ dilation was associated with elevated CSF cAMP (1037±58 vs. 1919±209 fmol/ml for control and 10 −6 M NOC/oFQ). Glibenclamide and iberiotoxin, K ATP and K ca channel antagonists, attenuated NOC/oFQ induced dilation (15±1 and 28±1 vs. 10±1 and 19±1% before and after iberiotoxin). In contrast, the nitric oxide synthase inhibitor, l-NNA, and the protein kinase G inhibitor, Rp 8-Br cGMPs had no effect on NOC/oFQ dilation while such dilation was not associated with a change in CSF cGMP. The putative ORL-1 receptor antagonist [F/G] NOC/oFQ (1–13)–NH 2 blocked NOC/oFQ dilation while responses were unchanged after naloxone (17±1 and 30±2 vs. 3±1 and 5±1%, before and after [F/G] NOC/oFQ (1–13)–NH 2). Dilation to other opioids (e.g., methionine enkephalin) was unchanged by [F/G] NOC/oFQ (1–13)–NH 2. These data show that NOC/oFQ elicits pial artery dilation, at least in part, via cAMP, K ATP, and K ca channel dependent mechanisms. These data suggest that such a mechanism involves the sequential release of cAMP and subsequent K ATP and K ca channel activation.

VEGF increases permeability of the blood-brain barrier via a nitric oxide synthase/cGMP-dependent pathway.

It appears that the expression of vascular endothelial growth factor (VEGF) is increased during brain injury and thus may contribute to disruption of the blood-brain barrier (BBB) during cerebrovascular trauma. The first goal of this study was to determine the effect of VEGF on permeability of the BBB in vivo. The second goal was to determine possible cellular mechanisms by which VEGF increases permeability of the BBB. We examined the pial microcirculation in rats using intravital fluorescence microscopy. Permeability of the BBB [clearance of FITC-labeled dextran of molecular mass 10,000 Da (FITC-dextran-10K)] and diameter of pial arterioles were measured in absence and presence of VEGF (0.01 and 0.1 nM). During superfusion with vehicle (saline), clearance of FITC-dextran-10K from pial vessels was minimal and diameter of pial arterioles remained constant. Topical application of VEGF (0.01 nM) did not alter permeability of the BBB to FITC-dextran-10K or arteriolar diameter. However, superfusion with VEGF (0.1 nM) produced a marked increase in clearance of FITC-dextran-10K and a modest dilatation of pial arterioles. To determine a potential role for nitric oxide and stimulation of soluble guanylate cyclase in VEGF-induced increases in permeability of the BBB and arteriolar dilatation, we examined the effects of NG-monomethyl-L-arginine (L-NMMA; 10 microM) and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 1.0 microM), respectively. L-NMMA and ODQ inhibited VEGF-induced increases in permeability of the BBB and arteriolar dilatation. The findings of the present study suggest that VEGF, which appears to be increased in brain tissue during cerebrovascular trauma, increases the permeability of the BBB via the synthesis/release of nitric oxide and subsequent activation of soluble guanylate cyclase.

Carbon monoxide and cerebral microvascular tone in newborn pigs.

The present study addresses the hypothesis that CO produced from endogenous heme oxygenase (HO) can dilate newborn cerebral arterioles. HO-2 protein was highly expressed in large and small blood vessels, as well as parenchyma, of newborn pig cerebrum. Topically applied CO dose-dependently dilated piglet pial arterioles in vivo over the range 10(-11)-10(-9) M (maximal response). CO-induced cerebrovascular dilation was abolished by treatment with the Ca2+-activated K+ channel inhibitors tetraethylammonium chloride and iberiotoxin. The HO substrate heme-L-lysinate also produced tetraethylammonium-inhibitable, dose-dependent dilation from 5 x 10(-8) to 5 x 10(-7) M (maximal). The HO inhibitor chromium mesoporphyrin blocked dilation of pial arterioles in response to heme-L-lysinate. In addition to inhibiting dilation to heme-L-lysinate, chromium mesoporphyrin also blocked pial arteriolar dilations in response to hypoxia but did not alter responses to hypercapnia or isoproterenol. We conclude that CO dilates pial arterioles via activation of Ca2+-activated K+ channels and that endogenous HO-2 potentially can produce sufficient CO to produce the dilation.

Possible obligatory functions of cyclic nucleotides in hypercapnia-induced cerebral vasodilation in adult rats.

Current evidence suggests that nitric oxide (NO) and vasodilating prostanoids, possibly via the actions of cGMP and cAMP, play permissive roles in hypercapnic cerebral vasodilation. The present study examined whether cGMP and cAMP have obligatory functions in hypercapnia. Using a closed cranial window in adult rats, we measured pial arteriolar diameters and periarachnoid cerebrospinal fluid (pCSF) cyclic nucleotide levels during normo- and hypercapnia and in the presence or absence of inhibitors of neuronal NO synthase (nNOS) or cyclooxygenase (COX). Also, we measured cGMP and cAMP contents in primary neuronal and astrocyte cultures, at different levels of CO2. Hypercapnia (arterial PCO2 65 mmHg)-induced pial arteriolar dilation was accompanied by 70-80% elevations in pCSF cGMP and cAMP. Inhibition of nNOS with 7-nitroindazole (7-NI) significantly reduced both the CO2-induced arteriolar dilation (by 77%) and the pCSF cGMP and cAMP increases (by 60-70%). Inhibition of COX with indomethacin reduced arteriolar CO2 reactivity (by 83%) and pCSF cyclic nucleotide increases (by 80-100%). In neuronal cultures a transient NO-dependent increase in cGMP, but not cAMP, was seen when the CO2 level was raised from 5 to 14%. No changes were seen in astrocytes. The 7-NI and indomethacin-inhibitable increases in pial arteriolar diameter and cyclic nucleotide production during hypercapnia suggest a link between these two responses. One possible, although not exclusive, interpretation of these findings is that the cyclic nucleotides have an obligatory function in the CO2 response. The large overlap in the abilities of nNOS and COX inhibitors to elicit those effects further implies interactions ("cross talk") between the cGMP and cAMP vasodilating pathways. The in vitro data suggest that hypercapnia stimulates NO production in neurons.

Loss of endothelium and receptor-mediated dilation in pial arterioles of rats fed a short-term high salt diet.

A high salt diet often is regarded as an accessory risk factor in hypertension, coincidental to the deleterious effect of high blood pressure on vasodilator function. The aim of this study was to determine whether short-term ingestion of a high salt diet per se impairs vasodilator function in the cerebral circulation independent of blood pressure changes. Adult Sprague-Dawley rats were fed a normal salt (0.8%) or high salt (4%) diet for 3 days. Mean arterial pressures were similar in the normal and high salt groups (123+/-2 and 125+/-2 mm Hg, respectively). Subsequently, the responses of the in situ pial arterioles to acetylcholine, iloprost, and sodium nitroprusside were determined in cranial windows using intravital videomicroscopy. Pial arterioles of rats fed normal and high salt diets showed similar resting diameters of 69+/-2 and 72+/-3 microm, respectively, but their reactivity patterns to vasodilator stimuli were markedly different. Arterioles of rats fed a normal salt diet dilated progressively up to 17+/-3% in response to the endothelium-dependent agent acetylcholine (10(-9) to 10(-6) mol/L) and dilated by 22+/-2% in response to the prostaglandin I2 receptor agonist iloprost (3x10(-11) mol/L). In contrast, pial arterioles of rats fed a high salt diet constricted by 4+/-3% and 8+/-2% in response to acetylcholine and iloprost, respectively. Sodium nitroprusside (10(-6) mol/L), a nitric oxide donor, dilated pial arterioles of rats fed low and high salt diets by a similar amount (19+/-3% and 16+/-2%, respectively), suggesting that signaling mechanisms for dilation distal to the vascular smooth muscle membrane were intact after high salt intake. These results provide the first evidence that the short-term ingestion of a high salt diet may severely impair the vasodilator function of the in situ cerebral microcirculation independent of blood pressure elevation.

Role of tyrosine phosphorylation in the regulation of cerebral vascular tone in newborn pig in vivo.

The role of tyrosine phosphorylation was investigated using protein tyrosine phosphatase inhibitors in newborn pigs equipped with a cranial window in vivo. We tested the hypothesis that cyclooxygenase and nitric oxide (NO) synthase are physiological targets for tyrosine phosphorylation in cerebral circulation. Phenylarsine oxide dilated pial arterioles and increased prostacyclin and prostaglandin E2 in cortical periarachnoid fluid; these responses were inhibited by indomethacin. Nomega-nitro-L-arginine methyl ester (L-NAME) and Nomega-nitro-L-arginine (L-NNA) inhibited the vasodilation to phenylarsine oxide; the effects of NO synthase inhibitors and indomethacin were additive. Cyclooxygenase-mediated vascular responses were assessed using topical application of arachidonic acid. Phenylarsine oxide and sodium orthovanadata potentiated vasodilation and prostanoid synthesis in response to arachidonic acid. Nomega-nitro-L-arginine methyl ester and Nomega-nitrol-arginine did not affect vasodilation or prostanoid production in response to arachidonic acid, indicating no cross talk between cyclooxygenase and NO synthase. These data indicate that cyclooxygenase and NO synthase are physiological targets for tyrosine phosphorylation in the cerebral circulation of newborn pigs.

PTX-sensitive G proteins and permissive action of prostacyclin in newborn pig cerebral circulation.

The present study of newborn pig cerebral circulation investigated the role of pertussis toxin (PTX)-sensitive GTP binding proteins in the permissive action of prostacyclin in specific dilator responses. Pial arterioles of anesthetized piglets were observed through closed cranial windows. The piglets were treated topically with PTX and intravenously with indomethacin. The effects of hypercapnia (10% CO2 ventilation) and topical 5,6-epoxyeicosatrienoic acid (5,6-EET) on pial arteriolar diameter were noted before and after the intervention. Samples of the artificial cerebrospinal fluid (aCSF) were collected from beneath the cranial windows for determination of the cAMP concentration. After administration of PTX, indomethacin still abolished pial arteriolar dilation to both hypercapnia and 5, 6-EET and also inhibited the cAMP elevation caused by hypercapnia. The addition of phorbol 12-myristate 13-acetate (PMA), but not iloprost, restored the increase in cAMP and vascular responses to hypercapnia and 5,6-EET. Therefore, in the newborn pig cerebral microvasculature, PTX appears to inhibit a G protein involved in the permissive action of prostacyclin. However, the protein kinase C (PKC) activator PMA appears to act downstream from the block, and, therefore, the permissive action of PMA is not affected by PTX. We suggest that the prostacyclin IP receptor may be coupled to phospholipase C via a PTX-sensitive G protein that normally permits vasodilation to specific stimuli via activation of a PKC, resulting in phosphorylation of a component of the adenylyl cyclase pathway.

Effect of lipopolysaccharide on the permeability and reactivity of the cerebral microcirculation: role of inducible nitric oxide synthase

The goal of this study was to examine the effect of lipopolysaccharide on the permeability of the blood–brain barrier and reactivity of cerebral arterioles. We examined the pial microcirculation in rats using intravital fluorescence microscopy. Permeability of the blood–brain barrier (clearance of fluorescent-labeled dextran; molecular weight 10,000 Da; FITC-dextran-10K) and diameter of pial arterioles were measured in the absence and presence of topical application of vehicle (saline) or lipopolysaccharide (200 ng/ml). During superfusion with vehicle, clearance of FITC-dextran-10K from pial vessels was minimal, and diameter of pial arterioles remained constant. Topical application of lipopolysaccharide (200 ng/ml) produced an increase in clearance of FITC-dextran-10K and dilated pial arterioles. To determine whether lipopolysaccharide-induced changes in permeability of the blood–brain barrier and dilatation of cerebral arterioles was related to the synthesis/release of inducible nitric oxide, we examined the effects of aminoguanidine (0.5 mM). Aminoguanidine inhibited lipopolysaccharide-induced increases in permeability of the blood–brain barrier and dilatation of cerebral arterioles. The findings of the present study suggest that lipopolysaccharide increases permeability of the blood–brain barrier and diameter of pial arterioles via the activation of inducible nitric oxide synthase.

Mild hypothermia can attenuate nitroglycerin-induced vasodilation of pial arterioles in the cat.

The purpose of the present study was to investigate the effect of mild hypothermia on nitroglycerin-induced vasodilation of cerebral vessels. The cranial window technique, combined with microscopic video recording, was used in an experiment involving 26 cats anesthetized with isoflurane. Animals were randomly assigned to either a normothermic or a mildly hypothermic group (33 degrees C). We administered three different concentrations of nitroglycerin (10[-6], 10[-5], 10[-4] M) under the window and measured the diameter of small (< 100 microm) and large (100-200 microm) pial arterioles. In the normothermic group (n = 13), nitroglycerin produced a significant dilation of both small and large arterioles in a dose-dependent manner. In the hypothermic group (n = 13), a significant dilation of arterioles was observed only after topical application of nitroglycerin at a concentration of 10(-4) M. The percent increase in diameter of small and large arterioles was less in the hypothermic group than the normothermic group. Our in vivo study demonstrates that topically applied nitroglycerin produces a dose-dependent dilation of pial arterioles in normothermic cats anesthetized with isoflurane, but the reduction of temperature to 33 degrees C significantly attenuates nitroglycerin-induced vasodilation of pial arterioles. Although nitroglycerin may be used in hypothermic patients, the effect of mild hypothermia on nitroglycerin-induced vasodilation of cerebral vessels is unknown. In this study, we investigated the effects of nitroglycerin on pial arteriolar diameter in normothermic and hyperthermic cats. Hypothermia was found to attenuate nitroglycerin-induced vasodilation.

Mild Hypothermia Can Attenuate Nitroglycerin-Induced Vasodilation of Pial Arterioles in the Cat

The purpose of the present study was to investigate the effect of mild hypothermia on nitroglycerin-induced vasodilation of cerebral vessels. The cranial window technique, combined with microscopic video recording, was used in an experiment involving 26 cats anesthetized with isoflurane. Animals were randomly assigned to either a normothermic or a mildly hypothermic group (33[degree sign]C). We administered three different concentrations of nitroglycerin (10-6, 10-5, 10-4 M) under the window and measured the diameter of small (<100 [micro sign]m) and large (100-200 [micro sign]m) pial arterioles. In the normothermic group (n = 13), nitroglycerin produced a significant dilation of both small and large arterioles in a dose-dependent manner. In the hypothermic group (n = 13), a significant dilation of arterioles was observed only after topical application of nitroglycerin at a concentration of 10-4 M. The percent increase in diameter of small and large arterioles was less in the hypothermic group than the normothermic group. Our in vivo study demonstrates that topically applied nitroglycerin produces a dose-dependent dilation of pial arterioles in normothermic cats anesthetized with isoflurane, but the reduction of temperature to 33[degree sign]C significantly attenuates nitroglycerin-induced vasodilation of pial arterioles. Implications: Although nitroglycerin may be used in hypothermic patients, the effect of mild hypothermia on nitroglycerin-induced vasodilation of cerebral vessels is unknown. In this study, we investigated the effects of nitroglycerin on pial arteriolar diameter in normothermic and hyperthermic cats. Hypothermia was found to attenuate nitroglycerin-induced vasodilation. (Anesth Analg 1998;86:546-51)