Perivascular Sympathetic Nerves Research Articles

Human Orbital Sympathetic Nerve Pathways

To determine pathways of sympathetic nerves from the orbital apex to the eyelids in human cadaver tissue using immunohistochemistry. Human cadaver orbit tissue was sectioned and immunolabeled with a monoclonal antityrosine hydroxylase antibody. In the orbital apex, the nasociliary, frontal, lacrimal, and maxillary branches of the trigeminal nerve demonstrated intense staining upon entering the orbit. Immunoreactive axons from the nasociliary and frontal nerves were observed to join the extraocular motor nerves in the posterior orbit. A plexus of immunolabeled nerves was observed to accompany the ophthalmic artery as it entered the orbital apex. The ophthalmic artery and its branches throughout the orbit demonstrated staining of nerve fibers in the peripheral muscularis. The nasociliary nerve contributed sympathetic branches to the ciliary ganglion. Nerves passing through the ciliary ganglion and a few ganglion cell bodies demonstrated mild to moderate tyrosine hydroxylase reactivity. Axons within the short and long ciliary nerves demonstrated strong tyrosine hydroxylase reactivity and were observed to enter the posterior sclera and the suprachoroidal space. The lacrimal gland demonstrated mild pericapillary staining and occasional stromal nerve fibers reactive to the antityrosine hydroxylase antibody. Müller muscle and the inferior tarsal muscle possessed a strong tyrosine hydroxylase-reactive nerve supply that appeared to originate from the anterior terminal branches of the nasociliary and lacrimal nerves. Sympathetic nerves enter the orbit via the first and second divisions of the trigeminal nerve and a plexus of nerves surrounding the ophthalmic artery. Extraocular motor nerves receive a sympathetic nerve supply from the sensory nerves in the posterior orbit. Some ciliary ganglion cell bodies demonstrated tyrosine hydroxylase-like reactivity, suggesting a sympathetic modulatory role for the ciliary ganglion. Sympathetics innervate ocular structures via the posterior ciliary nerves. Sympathetic axons travel anteriorly in the orbit via the nasociliary and lacrimal nerves to innervate the sympathetic eyelid muscles. Sympathetic nerves also travel with the frontal branch of the ophthalmic nerve to innervate the forehead skin. The ophthalmic artery and all of its branches contain a perivascular sympathetic nerve supply that may be involved in regulation of blood flow to ocular and orbital structures.

The Influence of Pregnancy and Gender on Perivascular Innervation of Rat Posterior Cerebral Arteries

The authors investigated the influence of pregnancy and gender on the density of trigeminal and sympathetic perivascular nerves in posterior cerebral arteries (PCA) and the reactivity to norepinephrine and calcitonin gene-related peptide (CGRP). PCAs were isolated from nonpregnant, late-pregnant, postpartum, and male rats, mounted and pressurized on an arteriograph chamber to obtain concentration-response curves to norepinephrine and CGRP. Arteries were immunostained for CGRP-, tyrosine hydroxylase-, and protein gene product 9.5 (PGP 9.5)-containing perivascular nerves, and nerve density was determined morphologically. Pregnancy had a trophic effect on trigeminal perivascular innervation (P < .01 vs male); however, this was not accompanied by a change in reactivity to CGRP. Sympathetic and PGP 9.5 nerve densities were not altered by pregnancy or gender, and there were no differences in reactivity to norepinephrine. Together, these results suggest that the increase in trigeminal innervation during pregnancy is more related to nociception than in controlling resting cerebral blood flow.

Differences in sympathetic neuroeffector transmission to rat mesenteric arteries and veins as probed by in vitro continuous amperometry and video imaging

As arteries are resistance blood vessels while veins perform a capacitance function, it might be expected that sympathetic neural control of arteries and veins would differ. The function of sympathetic nerves supplying mesenteric arteries (MA) and veins (MV) in rats was investigated using in vitro continuous amperometry with a carbon fibre microelectrode and video imaging. We simultaneously measured noradrenaline (NA) overflow at the blood vessel adventitial surface and vasoconstriction evoked by electrical stimulation of perivascular sympathetic nerves. Sympathetic nerve arrangement was studied using glyoxylic acid-induced fluorescence of NA. We found that: (i) there were significant differences between MA and MV in the arrangement of sympathetic nerves; (ii) frequency-response curves for NA overflow and vasoconstriction for MV were left-shifted compared to MA; (iii) the P2X receptor antagonist, pyridoxal-phosphate-6-azophenyl-2',4'-disulphonic acid (PPADS, 10 microm), reduced constrictions in MA but not in MV while the alpha(1)-adrenergic receptor antagonist, prazosin (0.1 microm), blocked constrictions in MV but not in MA; (iv) NA overflow for MA was enhanced by the alpha(2)-adrenergic receptor antagonist, yohimbine (1.0 microm), and attenuated by the alpha(2)-adrenergic receptor agonist, UK 14,304 (1.0 microm), while yohimbine and UK 14,304 had little effect in MV; (v) cocaine (10 microm) produced larger increases in NA overflow in MA than in MV; (vi) UK 14,304 constricted MV but not MA while yohimbine reduced constrictions in MV but not MA. We conclude that there are fundamental differences in sympathetic neuroeffector mechanisms in MA and MV, which are likely to contribute to their different haemodynamic functions.

On the presence of neurotrophin p75 receptor on rat sympathetic cerebrovascular nerves

Although the presence of neurotrophin p75 receptor on sympathetic nerves is a well-recognised feature, there is still a scarcity of details of the distribution of the receptor on cerebrovascular nerves. This study examined the distribution of p75 receptor on perivascular sympathetic nerves of the middle cerebral artery and the basilar artery of healthy young rats using immunohistochemical methods at the laser confocal microscope and transmission electron microscope levels. Immunofluorescence methods of detection of tyrosine hydroxylase (TH) in sympathetic nerves, p75 receptor associated with the nerves, and also S-100 protein in Schwann cells were applied in conjunction with confocal microscopy, while the pre-embedding single and double immunolabelling methods (ExtrAvidin and immuno-gold-silver) were applied for the electron microscopic examination. Immunofluorescence studies revealed "punctuate" distribution of the p75 receptor on sympathetic nerves including accompanying Schwann cells. Image analysis of the nerves showed that the level of co-localization of p75 receptor and TH was low. Immunolabelling applied at the electron microscope level also showed scarce co-localization of TH (which was intra-axonal) and p75. Immunoreactivity for p75 receptor was present on the cell membrane of perivascular axons and to a greater extent on the processes of accompanying Schwann cells. Some Schwann cell processes were adjacent to each other displaying strong immunoreactivity for p75 receptor; immunoreactivity was located on the extracellular sites of the adjacent cell membranes suggesting that the receptor was involved in cross talk between these. It is likely that variability of locations of p75 receptor detected in the study reflects diverse interactions of p75 receptor with axons and Schwann cells. It might also imply a diverse role for the receptor and/or the plasticity of sympathetic cerebrovascular nerves to neurotrophin signalling.

Distribution of the Tissue Plasminogen Activator (t‐PA) Promotor to Perivascular Sympathetic Nerves and Other Neural Crest Derivatives: Evidence of a Dispersed Plasmin Proteolysis

Distribution of the Tissue Plasminogen Activator (t‐PA) Promotor to Perivascular Sympathetic Nerves and Other Neural Crest Derivatives: Evidence of a Dispersed Plasmin Proteolysis

Sympathetically evoked Ca2+signaling in arterial smooth muscle

The sympathetic nervous system plays an essential role in the control of total peripheral vascular resistance and blood flow, by controlling the contraction of small arteries. Perivascular sympathetic nerves release ATP, norepinephrine (NE) and neuropeptide Y. This review summarizes our knowledge of the intracellular Ca2+ signals that are activated by ATP and NE, acting respectively on P2X1 and alpha1-adrenoceptors in arterial smooth muscle. Each neurotransmitter produces a unique type of post-synaptic Ca2+ signal and associated contraction. The neural release of ATP and NE is thought to vary markedly with the pattern of nerve activity, probably reflecting both pre- and post-synaptic mechanisms. Finally, we show that Ca2+ signaling during neurogenic contractions activated by trains of sympathetic nerve fiber action potentials are in fact significantly different from that elicited by simple bath application of exogenous neurotransmitters to isolated arteries (a common experimental technique), and end by identifying important questions remaining in our understanding of sympathetic neurotransmission and the physiological regulation of contraction of small arteries.

Novel sites of aldose reductase immunolocalization in normal and streptozotocin‐diabetic rats

Glucose metabolism by aldose reductase (AR) is implicated in the pathogenesis of many diabetic complications, including neuropathy. We have re-evaluated the distribution of AR in the sciatic nerve and dorsal root ganglion (DRG) of normal rats, expanded these observations to describe the location of AR in the spinal cord and footpad skin, and investigated whether diabetes alters the distribution of AR. In sciatic nerve, AR was restricted to cytoplasm of myelinated Schwann cells and endothelial cells of epineurial, but not endoneurial, blood vessels. AR immunoreactivity (IR) was present in satellite cells in the DRG. In skin, AR-IR was detected in vascular endothelial cells, Schwann cells of myelinated fibers, and axons of perivascular sympathetic nerves. AR was localized selectively to oligodendrocytes of the white matter of spinal cord. The distribution of AR-IR in sciatic nerve, DRG, skin, and spinal cord was not altered by up to 12 weeks of streptozotocin-induced diabetes. Identification of perineuronal satellite cells, oligodendrocytes, and perivascular sympathetic nerves as AR-expressing cells reveals them as cellular sites with the potential to contribute to diabetic neuropathy.

P2X1 receptors mediate sympathetic postjunctional Ca2+ transients in mesenteric small arteries

Brief, spatially localized Ca(2+) transients occur in the smooth muscle adjacent to perivascular nerves of small arteries during neurogenic contractions. We named these "junctional Ca(2+) transients" (jCaTs) and postulated that they arose from Ca(2+) entering smooth muscle cells through P2X(1) receptors activated by neurally released ATP. Nevertheless, the lack of potent, subtype-selective P2X-receptor antagonists made determining the exact molecular identity of the channels difficult. Here we used small, pressurized mesenteric arteries from P2X(1)-receptor-deficient mice (KO) to test the hypothesis that jCaTs arise from Ca(2+) entering the smooth muscle cell via P2X(1) receptors. In wild-type (WT) arteries, confocal microscopy of fluo-4 fluorescence during electrical field stimulation (EFS) of perivascular sympathetic nerves revealed jCaTs in the smooth muscle cells adjacent to the perivascular nerves, similar to those reported previously in rat arteries, and alpha-latrotoxin (2.5 nM) markedly increased the frequency of "spontaneous" jCaTs. In the KO arteries, however, neither EFS nor alpha-latrotoxin elicited any jCaTs. A potent P2X-receptor agonist, alpha,beta-methylene ATP (10.0 microM), elicited strong contractions and increased intracellular Ca(2+) concentration in WT arteries but elicited neither in KO arteries. A biphasic vasoconstriction in response to EFS was observed in WT arteries. In KO arteries, however, the initial rapid, transient component of the biphasic vasoconstriction was absent. The data support the hypothesis that jCaTs represent Ca(2+) that enters the smooth muscle cells through P2X(1) receptors activated by neurally released ATP and that this Ca(2+) is involved in the initial rapid component of the sympathetic neurogenic contraction.

Novel localization of CD38 in perivascular sympathetic nerve terminals

Using high performance liquid chromatography fraction analysis we have recently established that numerous smooth muscle preparations, including the canine mesenteric artery and vein, release β-nicotinamide adenine dinucleotide upon short-pulse electrical field stimulation in tetrodotoxin- and ω-conotoxin GVIA-sensitive manners [ Smyth LM, Bobalova J, Mendoza MG, Lew C, Mutafova-Yambolieva VN (2004) Release of beta-nicotinamide adenine dinucleotide upon stimulation of postganglionic nerve terminals in blood vessels and urinary bladder. J Biol Chem 279:48893–48903.]. The β-nicotinamide adenine dinucleotide metabolites ADP-ribose and cyclic ADP-ribose are also present in the tissue superfusates. CD38 is a multifunctional enzyme involved in the degradation of β-nicotinamide adenine dinucleotide to ADP-ribose and cyclic ADP-ribose. Western immunoblot analysis revealed that CD38 is expressed in both artery and vein. Confocal laser scanning microscopy established colocalization of CD38 with tyrosine hydroxylase, synaptotagmin and synaptic vesicle protein in both blood vessels. High performance liquid chromatography with fluorescence detection demonstrated that whole tissue segments metabolize 1, N 6-etheno-nicotinamide adenine dinucleotide to 1, N 6-etheno-ADP-ribose and nicotinamide-guanine dinucleotide to cyclic GDP-ribose, suggesting the presence of both nicotinamide adenine dinucleotide-glycohydrolase and ADP-ribosyl cyclase activities in these blood vessels. Both enzymes appear to be associated with the membrane fraction, and therefore might be attributed to CD38. These data demonstrate a previously uncharacterized localization of CD38 in perivascular autonomic nerve terminals. Therefore, the β-nicotinamide adenine dinucleotide/CD38 system may provide new mechanisms in autonomic neurovascular control.

Sympathetic neural inhibition of conducted vasodilatation along hamster feed arteries: complementary effects of α1‐ and α2‐adrenoreceptor activation

Vasodilatation initiated on arterioles of skeletal muscle ascends into the proximal feed arteries through cell-to-cell conduction along the endothelium and into smooth muscle. Whereas perivascular sympathetic nerve activity (SNA) can inhibit conducted vasodilatation and restrict muscle blood flow, the signalling events mediating this interaction are poorly defined. Therefore, using isolated pressurized (75 mmHg) feed arteries (diameter (microm) at rest = 53 +/- 3; maximum = 99 +/- 2; n = 86) of the hamster retractor muscle, we tested the hypothesis that distinct yet complementary signalling pathways underlie the ability of SNA to inhibit conduction. Conducted vasodilatation was initiated using ACh microiontophoresis (1 microA; 250, 500 and 1000 ms) and SNA was initiated using local field stimulation (30-50 V; 1 ms at 2, 8 and 16 Hz). With vasodilatations of 5-20 microM, conduction increased with ACh pulse duration and was inhibited progressively as the frequency of SNA increased. During SNA, conduction was partially restored with inhibition of alpha1- (0.1 microM prazosin) or alpha2- (0.1 microM RX821002) adrenoreceptors and fully restored with both antagonists present. Activating alpha1- (50 nM phenylephrine) or alpha2- (1 microM UK 14,304) adrenoreceptors inhibited conduction partially and their simultaneous activation inhibited conduction cumulatively (P < 0.05). Elevated [K+]o (30 or 40 mM) or phorbol esters (0.5 microM) also inhibited conduction yet similar constriction with l-NNA (50 microM) or Bay K 8644 (10 nM) did not. Thus, the activation of alpha1- and alpha2-adrenoreceptors inhibits conducted vasodilatation through complementary signalling events. With robust coupling along the endothelium, our modelling predicts that the inhibition of conduction by SNA can be explained by reduced electrical coupling through myoendothelial gap junctions or greater current leak across smooth muscle cell membranes.

Unusual absence of endothelium-dependent or -independent vasodilatation to purines or pyrimidines in the rat renal artery

Adenosine triphosphate (ATP) is a cotransmitter with noradrenaline (NA) in sympathetic perivascular nerves. It has a dual role in the maintenance of vascular tone as ATP, released from endothelial cells during shear stress or hypoxia, induces vasodilatation via endothelial P2Y receptors or by direct action on smooth muscle. The role and distribution of P2 receptors is well characterized for many blood vessels but not for the rat renal artery. This study aims to determine whether ATP is a vasoconstrictor cotransmitter with NA and whether ATP induces vasodilatation via the endothelium or smooth muscle. On isolated rat renal arteries, electrical field stimulation (EFS) in the absence and presence of antagonists to P2X receptors and alpha1-adrenoceptors was examined. Concentration-response curves were constructed to NA, ATP, alpha,beta-methylene ATP (alpha,beta-meATP), uridine triphosphate (UTP), and 2-methylthio ADP (2-MeSADP) on low tone. Curves to acetylcholine (ACh), 2-MeSADP, and UTP were constructed on raised tone. Immunofluorescent localization of P2X and P2Y receptor subtypes was performed. Electrical field stimulation induced vasoconstriction, partially inhibited by the P2X receptor antagonist, pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid, and predominantly by prazosin. Exogenous NA and ATP mimicked EFS; immunostaining for P2X1 and P2X2 receptors was expressed on vascular smooth muscle. Unusually, ATP, 2-MeSADP, and UTP failed to induce vasodilatation. Acetylcholine induced vasodilatation. alpha,beta-meATP, 2-MeSADP, and UTP induced vasoconstriction via P2X1, P2Y1, and P2Y2 receptors, respectively. Immunostaining for P2X1, P2Y1, and P2Y2 receptors was expressed on the vascular smooth muscle. Adenosine triphosphate and NA are cotransmitters in sympathetic nerves supplying the rat renal artery, NA being the dominant partner. The novel feature of this vessel is that purines and pyrimidines do not produce either endothelium-dependent or -independent vasodilatation; P2X1, P2Y1, and P2Y2 receptors on the smooth muscle all mediate vasoconstriction.

Sympathetic nerves inhibit conducted vasodilatation along feed arteries during passive stretch of hamster skeletal muscle.

Ascending vasodilatation is integral to blood flow control in exercising skeletal muscle and is attributable to conduction from intramuscular arterioles into proximal feed arteries. Passive stretch of skeletal muscle can impair muscle blood flow but the mechanism is not well understood. We hypothesized that the conduction of vasodilatation along feed arteries can be modulated by changes in muscle length. In anaesthetized hamsters, acetylcholine (ACh) microiontophoresis triggered conducted vasodilatation along feed arteries (diameter, 50-70 microm) of the retractor muscle secured at 100 % resting length or stretched by 30 %. At 100 % length, ACh evoked local dilatation (> 30 microm) and this response conducted rapidly along the feed artery (14 +/- 1 microm dilatation at 1600 microm upstream). During muscle stretch, feed arteries constricted approximately 10 microm (P < 0.05) and local vasodilatation to ACh was maintained while conducted vasodilatation was reduced by half (P < 0.01). Resting diameter and conduction recovered upon restoring 100 % length. Sympathetic nerve stimulation (4-8 Hz) produced vasoconstriction and attenuated conduction in the manner observed during muscle stretch, as did noradrenaline or phenylephrine (10 nM). Inhibiting nitric oxide production (Nomega-nitro-L-arginine, 50 microM) produced similar vasoconstriction yet had no effect on conduction. Phentolamine, prazosin, or tetrodotoxin (1 microM) during muscle stretch abolished vasoconstriction and restored conduction. Inactivation of sensory nerves with capsaicin had no effect on vasomotor responses. Thus, muscle stretch can attenuate conducted vasodilatation by activating alpha-adrenoreceptors on feed arteries through noradrenaline released from perivascular sympathetic nerves. This autonomic feedback mechanism can restrict muscle blood flow during passive stretch.

Electrophysiological effects of activating the peptidergic primary afferent innervation of rat mesenteric arteries.

1. Intracellular recording was used to investigate the electrophysiological effects of activating peptidergic primary afferent axons with capsaicin in the smooth muscle of rat mesenteric arteries in vitro. In addition, continuous amperometry was used to monitor the effects of capsaicin on noradrenaline release from the sympathetic nerves. 2. Capsaicin (1 microm) produced a hyperpolarization (-11+/-2 mV) and a reduction in the time constant of decay of excitatory junction potentials (e.j.p.'s) evoked by electrical stimulation of the perivascular sympathetic nerves. These effects of capsaicin were mimicked by calcitonin gene-related peptide (CGRP; 1 and 10 nm) but not by substance P (50 nm), which produced a small hyperpolarization (maximum -3+/-1 mV) but did not change excitatory junction potential (e.j.p.) time course. 3. The hyperpolarization produced by capsaicin and CGRP was blocked by glibenclamide (10 microm) but was not changed by the CGRP antagonist, CGRP8-37 (0.5 microm). Mechanical denudation of the endothelium also did not reduce the effect of capsaicin on membrane potential. 4. Capsaicin (1 microm) increased the amplitude of e.j.p.'s. This effect was not mimicked by CGRP or substance P nor blocked by glibenclamide or CGRP8-37. 5. All effects of capsaicin desensitized. 6. Capsaicin (1 microm) had no effect on noradrenaline-induced oxidation currents evoked by electrical stimulation, indicating that noradrenaline release was unchanged. 7. These results suggest that CGRP released from primary afferent axons hyperpolarizes vascular smooth muscle by activating glibenclamide-sensitive K+ channels. The findings also indicate that an unknown factor released by the primary afferent axons increases e.j.p. amplitude.

Pb2+ inhibition of sympathetic alpha 7-nicotinic acetylcholine receptor-mediated nitrergic neurogenic dilation in porcine basilar arteries.

Chronic exposure to inorganic lead (Pb2+) has been shown to facilitate peripheral vasoconstriction causing hypertension. Effect of lead on cerebral vascular function has not been reported. We have suggested in isolated porcine cerebral arteries that alpha 7-nicotinic acetylcholine receptors (alpha 7-nAChRs) on perivascular sympathetic nerves mediate calcium influx in these neurons, resulting in release of norepinephrine. The released norepinephrine then acts on presynaptic beta2-adrenoceptors located on the neighboring nitrergic nerve terminals, causing nitric oxide (NO) release and vasodilation. Because Pb2+ has been shown to inhibit alpha 7-nAChR-mediated responses in the central nervous system, effects of Pb2+ on alpha 7-nAChR-mediated nitrergic neurogenic dilation in isolated porcine basilar arteries and calcium influx in cultured superior cervical ganglion (SCG) cells of the pig were examined using in vitro tissue bath and confocal microscopic techniques. The results indicated that Pb2+ (but not Cd2+, Zn2+, or Al3+) in a concentration-dependent manner blocked relaxation of endothelium-denuded basilar arterial rings induced by nicotine (100 microM) and choline (1 mM) without affecting relaxation induced by sodium nitroprusside or isoproterenol. Furthermore, significant calcium influx in cultured SCG cells induced by choline and nicotine was attenuated specifically by Pb2+ with IC50 values comparable with those from tissue bath study. These results provide evidence supporting that lead is a likely antagonist for alpha 7-nAChRs that are found on postganglionic sympathetic adrenergic nerve terminals of SCG origin. Furthermore, these results indicate that lead can attenuate dilation of cerebral arteries by blocking sympathetic nerve-mediated release of NO from the perivascular nitrergic nerves.

Alpha7-nicotinic acetylcholine receptors on cerebral perivascular sympathetic nerves mediate choline-induced nitrergic neurogenic vasodilation.

It has been suggested in isolated porcine cerebral arteries that stimulation by nicotine of alpha7-nicotinic acetylcholine receptors (alpha7-nAChRs) on sympathetic nerves, but not direct stimulation of parasympathetic nitrergic nerves, caused nitrergic neurogenic dilation. Direct evidence supporting this hypothesis has not been presented. The present study, which used in vitro tissue bath and confocal microscopy techniques, was designed to determine whether choline, a selective agonist for alpha7-nAChRs, induced sympathetic-dependent nitrergic dilation of porcine basilar arterial rings. Choline and several nAChR agonists induced exclusive relaxation of basilar arterial rings without endothelium. The relaxation was blocked by tetrodotoxin, nitro-L-arginine, guanethidine, and beta2-adrenoceptor antagonists. Furthermore, the relaxation was blocked by methyllycaconitine and alpha-bungarotoxin (preferential alpha7-nAChR antagonists) and mecamylamine but was not affected by dihydro-beta-erythroidine (a preferential alpha4-nAChR antagonist). Confocal microscopic study demonstrated that choline and nicotine induced significant calcium influx in cultured porcine superior cervical ganglionic cells but failed to affect calcium influx in cultured sphenopalatine ganglionic cells, providing direct evidence that choline and nicotine did not act directly on the parasympathetic nitrergic neurons. The increased calcium influx in superior cervical ganglionic cells was attenuated by alpha-bungarotoxin and methyllycaconitine but not by dihydro-beta-erythroidine. These results support our hypothesis that activation of alpha7-nAChRs on cerebral perivascular sympathetic nerves causes calcium influx and the release of norepinephrine, which then act on presynaptic beta2-adrenoceptors located on the neighboring nitrergic nerve terminals, resulting in NO release and vasodilation. Endogenous choline may play an important role in regulating cerebral sympathetic activity and vascular tone.

Mechanism of action of hypotensive prostaglandins in patients with essential hypertension.

Despite extensive investigation, the biological mechanisms causing essential hypertension (EHT) remain unclear. To clarify the means by which hypotensive prostaglandins (Hypo-PGs, mainly PGE1 and PGE2) act in patients with EHT, the interaction between intravenously infused Hypo-PGs and pressor substances such as an adrenergic neurotransmitter, noradrenaline (NA) and angiotensin II (AII) was examined both in patients with EHT and in perfused isolated rabbit ear artery preparations. In patients with EHT, Hypo-PGs were shown to reduce the pressor responses to intravenously infused NA or AII, although no significant difference was found between the pressor responses to NA under basal conditions and the responses during intravenous infusion of Hypo-PGs. Animal studies were undertaken to investigate the inhibitory action of Hypo-PGs on the vasoconstrictive responses to electrical stimulation of the perivascular sympathetic nerves (VSNS) and to exogenous NA at pre- and postjunctional sites in blood vessel walls. The suppressive action of Hypo-PGs on the response to VSNS was shown to be more potent than that to their action on the response to exogenous NA. Thus, it was concluded that the hypotensive action of intravenously infused Hypo-PGs in patients with EHT may be more dependent on prejunctional sites than on the postjunctional sites in the walls of blood vessels.

Pharmacology and Physiology of Perivascular Nerves Regulating Vascular Function: Role of Perivascular Sympathetic Nerves and Regional Differences in the Features of Sympathetic Innervation of the Vascular System

Maintenance of blood pressure is mostly dependent on sympathetic "tone", and the sympathetic nerve innervates the entire vascular bed, excepting the capillaries. Although norepinephrine (NE) is the principal neurotransmitter released upon sympathetic nerve stimulation, neuropeptide Y and ATP are cotransmitters in various vascular tissues. In addition, dopamine and epinephrine, as well as acetylcholine, have been shown to be sympathetic neurotransmitters in specific vasculatures. Transmitter NE release is modified by a number of endogenous substances including the transmitter itself. Chronic denervation of the preganglionic fiber induces an increase in NE release per pulse, indicating postganglionic neuronal supersensitivity. So far, three main adrenoceptor types have been shown, alpha1, alpha2 and beta, each of which is further divided into at least three subtypes, as well as the alpha1L-adrenoceptor, a phenotype of the cloned alpha1a-adrenoceptor, in the blood vessel. Thus, the response of vessels with different receptor types to a transmitter varies quantitatively and even qualitatively from one vessel to another. The remarkable diversity in the sympathetic innervation mechanism in the vascular system may play an important role in regional variations in the regulation of blood flow. The sympathetic nerve also exerts long-term trophic action on the blood vessel. In conclusion, the sympathetic nervous system plays an important role not only in the regulation of cardiovascular dynamics but in the maintenance of the vessel structure, as well.

Increased blood pressure responses in neuropeptide Y transgenic rats.

Considering the coexistence of neuropeptide Y (NPY) and norepinephrine in perivascular sympathetic nerves and the known vasoconstrictor cooperation of NPY with norepinephrine, we investigated the involvement of NPY in long-term control of cardiovascular functions using NPY transgenic (NPY-tg) rats. These rats were developed by injection of the rat (Sprague-Dawley) pronuclei with a 14.5-kb clone of the rat structural NPY gene. When compared with nontransgenic littermates, NPY concentrations were significantly increased in a number of cardiovascular tissues of NPY-tg hemizygotes. Direct basal mean arterial pressure and heart rate were not changed, but calculated total vascular resistance was significantly increased in NPY-tg subjects. Arterial pressure increases, in response to norepinephrine injection, were greater in the NPY-tg rats. Also, the hypotension and bradycardia in response to hemorrhage were significantly reduced in NPY-tg subjects. These results indicate that NPY, when expressed in increased amounts, potentiates the pressor effects of norepinephrine and contributes to maintaining blood pressure during hemorrhage, but it does not alter resting blood pressure. These transgenic rats will facilitate studies of the role of NPY signaling in cardiovascular regulation, particularly regarding its functional cooperation with norepinephrine.

Ultrastructure of NADPH diaphorase-positive nerve fibers and their terminals in the rat cerebral arterial system

To investigate how perivascular NO synthase (NOS)-containing nerves in the cerebral arterial system are involved in controlling the cerebral circulation, we observed the ultrastructure of NOS-containing nerve fibers and their terminals by means of nicotinamide adenine dinucleotide hydrogen phosphate-diaphorase (NADPH-d) histochemistry. We also observed the correlation between NADPH-d stained perivascular nerves and the perivascular sympathetic nerves, by means of double staining with NADPH-d histochemistry and tyrosine hydroxylase (TH) immunohistochemistry at the light microscopic level. NADPH-d-positive nerve fibers showed dense distribution mainly in the rostral portion of the circle of Willis and proximal portions of its main branches, where some of the NADPH-d-positive fibers coexisted with TH-positive fibers in a single nerve bundle. NADPH-d-positive nerve fibers were unmyelinated and had close contact with NADPH-d-negative myelinated and unmyelinated nerve fibers in a single nerve bundle, and NADPH-d-positive nerve terminals also existed closely with NADPH-d-negative nerve terminals. The number of NADPH-d-positive nerve terminals and their ratio to all other terminals were significantly higher in the rostral portion of the circle of Willis and the proximal portion of its branches, than the caudal portion of the circle of Willis and the distal portion of its branches. Nerve terminals were observed to locate within 250 nm from the basal lamina of arterial smooth muscle cells in the rostral portion of the circle of Willis and proximal portion of its branching arteries. The present observation confirmed that NOS-containing nerve fibers truely innervate the smooth muscle cells of the arterial wall in the circle of Willis and its main branches. Close contact between NADPH-d-positive and -negative nerve fibers and terminals in these arterial portions may indicate that NOS-containing perivascalar nerves may work to modulate the rest of the other perivascular nervous system, such as the sympathetic nerves, to regulate the homeostasis of the arterial tonus.

Enhancement of ATP release in hindlimb sympathetic perivascular nerve of the golden hamster during hibernation.

The present study investigated the effects of hibernation and hibernating body temperature (10 degrees C) on the relative changes that may occur in adrenergic and purinergic perivascular neurotransmission of the golden hamster. The hindlimb resistance vessels and the tibial artery of age-matched controls, cold exposed controls and hibernated hamsters were examined by pharmacological and electrophysiological techniques. At 34 degrees C, electrical field stimulation (EFS; supramaximal voltage, 0.5 ms; for 10 s) in all three groups evoked only twitch responses at 1-5 Hz, which were inhibited by piridoxal phosphate-6-azophenyl-2',4'-disulphonic acid (PPADS), a 2PX receptor antagonist. At 10-50 Hz the twitch responses were followed by sustained contractile responses, which were inhibited by prazosin, an alpha1-adrenoceptor antagonist. These responses were markedly enhanced at higher frequencies in hibernated tissues. At 10 degrees C, EFS evoked only the PPADS-sensitive transient responses in all the three groups, and this was markedly enhanced in hibernated tissues. At 34 degrees C, a single stimulus evoked a PPADS-sensitive excitatory junction potential (EJP) in all three groups but a train of pulses (e.g. approximately 0.5) evoked EJPs and prazosin-sensitive sustained depolarizations. These responses were markedly enhanced in hibernated cells. At 10 degrees C, either a single stimulus or a train of stimuli evoked only transient PPADS-sensitive EJPs, which were markedly enhanced in hibernated cells. The contractile responses and electrical membrane responses to exogenous ATP (1-1000 microM) and noradrenaline (0.1-100 microM) were unchanged in the three groups at 34 and at 10 degrees C. These results suggest that during hibernation enhancement of ATP release from the sympathetic perivascular nerves may occur, leading to an efficient means for maintenance of vascular tone and peripheral resistance.