Neuronal and non-neuronal modulation of sympathetic neurovascular transmission.

Noradrenaline, neuropeptide Y and adenosine triphosphate are co-stored in, and co-released from, sympathetic nerves. Each transmitter modulates its own release as well as the release of one another; thus, anything affecting the release of one of these transmitters has consequences for all. Neurotransmission at the sympathetic neurovascular junction is also modulated by non-sympathetic mediators such as angiotensin II, serotonin, histamine, endothelin and prostaglandins through the activation of specific pre-junctional receptors. In addition, nitric oxide (NO) has been identified as a modulator of sympathetic neuronal activity, both as a physiological antagonist against the vasoconstrictor actions of the sympathetic neurotransmitters, and also by directly affecting transmitter release. Here, we review the modulation of sympathetic neurovascular transmission by neuronal and non-neuronal mediators with an emphasis on the actions of NO. The consequences for co-transmission are also discussed, particularly in light of hypertensive states where NO availability is diminished.

A novel mechanism prevents the development of hypertension during chronic cold stress.

1 Chronic cold exposure of rats (7 days in a cold room at 4 degrees C) attenuated the sympathetic nerve stimulation (NS)-induced overflow of noradrenaline (NE) (measured by high-performance liquid chromatography, coupled to electrochemical detection) appearing in the perfusate/superfusate of the perfused mesenteric arterial bed as well as the increase in the perfusion pressure. 2 The same type of cold exposure resulted in an increase in tyrosine hydroxylase (TH) gene expression measured in the superior cervical ganglion and NE content measured in the mesenteric artery obtained from cold-exposed rats. 3 Addition of sodium nitroprusside, a nitric oxide (NO) donor, to the buffer perfusing the mesenteric arterial bed obtained from rats maintained at room temperature also resulted in an attenuation of the NS-induced overflow of NE and increase in perfusion pressure. 4 N(c)-nitro-L-arginine methyl ester (L-NAME), an NO synthase inhibitor, placed in the drinking water prevented the attenuation of the pre- and post-junctional responses to NS of the mesenteric arterial bed obtained from cold-exposed rats. 5 L-NAME treatment also increased the cold-induced elevation of blood pressure seen in whole animals. 6 The present results are consistent with the idea that cold exposure leads to a concomitant increase in sympathetic nerve activity and production of NO. We hypothesize that the increase in production and release of NO results in a decrease in the biologically active form of NE despite increased synthesis and release of the catecholamine. 7 It is concluded that the above-mentioned interactions serve as a protective mechanism offsetting the increased release and action of NE from sympathetic nerves and thus preventing the development of hypertension.

Endothelin (ET)-1-induced inhibition of ATP release from PC-12 cells is mediated by the ETB receptor: differential response to ET-1 on ATP, neuropeptide Y, and dopamine levels.

During sympathetic neurotransmitter release, there is evidence for differential modulation of cotransmitter release by endothelin (ET)-1. Using nerve growth factor (NGF)-differentiated PC12 cells, the effects of ET-1 on K(+)-stimulated release of ATP, dopamine (DA), and neuropeptide Y (NPY) were quantified using high-pressure liquid chromatography or radioimmunoassay. ET-1, in a concentration-dependent manner, inhibited the release of ATP, but not DA and NPY. Preincubation with the ET(A/B) antagonist, PD 142893 (N-acetyl-beta-phenyl-D-Phe-Leu-Asp-Ile-Ile-Trp), reversed the inhibitory effect of ET-1 on ATP release, which remained unaffected in the presence of the ET(A)-specific antagonist BQ123 [cyclo(D-Asp-Pro-D-Val-Leu-D-Trp)]. The ET(B) agonists, sarafotoxin 6c (Cys-Thr-Cys-Asn-Asp-Met-Thr-Asp-Glu-Glu-Cys-Leu-Asn-Phe-Cys-His-Gln-Asp-Val-Ile-Trp), BQ 3020 (N-acetyl-[Ala(11,15)]-endothelin 1 fragment 6-21Ac-Leu-Met-Asp-Lys-Glu-Ala-Val-Tyr-Phe-Ala-His-Leu-Asp-IIe-IIe-Trp), and IRL 1620 (N-succinyl-[Glu(9), Ala(11,15)]-endothelin 1 fragment 8-21Suc-Asp-Glu-Glu-Ala-Val-Tyr-Phe-Ala-His-Leu-Asp-Ile-Ile-Trp), decreased K(+)-stimulated release of ATP in a dose-dependent manner, and this effect was reversed by the ET(B) antagonists RES 701-1 [cyclic (Gly1-Asp9) (Gly-Asn-Trp-His-Gly-Thr-Ala-Pro-Asp-Trp-Phe-Phe-Asn-Tyr-Tyr-Trp)] and BQ 788 (N-[N-[N-[(2,6-dimethyl-1-piperidinyl)carbonyl]-4-methyl-l-leucyl]-1-(methoxycarbonyl)-D-tryptophyl]-D-norleucine sodium salt). Preincubation of PC12 cells with pertussis toxin reversed the ET-1-induced inhibition of the K(+)-evoked ATP release. Real-time intracellular calcium level recordings were performed on PC-12 cell suspensions, and ET-1 induced a dose-dependent decrease in the K(+)-evoked calcium levels. Nifedipine, the L-type voltage-dependent Ca(2+) channel antagonist, caused inhibition of the K(+)-stimulated ATP release, but the N-type Ca(2+) channel antagonist, omega-conotoxin GVIA, did not reverse the effect on ATP release. These data suggest that ET-1 modulates the release of ATP via the ET(B) receptor and its associated G(i/o) G-protein through attenuation of the influx of extracellular Ca(2+) through L-type channels.

Prostanoid-induced modulation of neuropeptide Y and noradrenaline release from the rat mesenteric bed.

1. A variety of prostanoids were examined for their ability to alter the periarterial nerve stimulation-induced release of noradrenaline (NA) and neuropeptide Y immunoreactive compounds (NPY-ir) from the perfused mesenteric arterial bed of the rat. 2. Periarterial nerve stimulation (16 Hz) increased the overflow of NA, NPY-ir and perfusion pressure. 3. The prostacyclin (PGI2) analogues, carbaPGI2 and cicaprost both produced a concentration-dependent attenuation of the nerve stimulation-induced increase in NA, NPY-ir overflow and perfusion pressure. 4. The prostaglandin (PG) analogue PGE2 attenuated the evoked increase in NPY-ir overflow as well as a modest decrease in NA. 5. PGE1, sulprostone and iloprost attenuated the nerve stimulation-induced increase in NA overflow but not NPY-ir. 6. Neither PGF2alpha nor the thromboxane A2 analogue U46619 altered the evoked increase in NA or NPY-ir overflow. 7. The results support the view that sympathetic co-transmitter release can be differentially modulated by paracrine/autocrine mediators at sympathetic neuroeffector junctions.

Catecholamine monoamine oxidase a metabolite in adrenergic neurons is cytotoxic in vivo.

3,4-Dihydroxyphenylglycolaldehyde is the monoamine oxidase-A metabolite of two catecholamine neurotransmitters, epinephrine and norepinephrine. This aldehyde metabolite and its synthesizing enzymes increase in cell bodies of catecholamine neurons in Alzheimer's disease. To test the hypothesis that 3,4-dihydroxyphenylglycolaldehyde, but not epinephrine or its major metabolite 4-hydroxy-3-methoxyphenylglycol, is a neurotoxin, we injected 3,4-dihydroxyphenylglycolaldehyde onto adrenergic neurons in the rostral ventrolateral medulla. Injections of epinephrine or 4-hydroxy-3-methoxyphenylglycol were made into the same area of controls. A dose response and time study were performed. Adrenergic neurons were identified by their content of the epinephrine synthesizing enzyme, phenylethanolamine N-methyltransferase, immunohistochemically. Apoptosis was evaluated by in situ terminal deoxynucleotidyl-transferase mediated dUTP nick end label staining. Injection of 3,4-dihydroxyphenylglycolaldehyde in amounts as low as 50 ng results in loss of adrenergic neurons and apoptosis after 18 h. The degree of neurotoxicity is dose and time dependent. Doses of 3,4-dihydroxyphenylglycolaldehyde 10-fold higher produce necrosis. Neither epinephrine nor 4-hydroxy-3-methoxyphenylglycol are toxic. A 2.5 microg injection of 3,4-dihydroxyphenylglycolaldehyde is toxic to cortical neurons but not glia. Active uptake of the catecholamine-derived aldehyde into differentiated PC-12 cells is demonstrated. Implications of these findings for catecholamine neuron death in neurodegenerative diseases are discussed.

Inactivation of catecholamines by superoxide gives new insights on the pathogenesis of septic shock.

A major feature of septic shock is the development of a vascular crisis characterized by nonresponsiveness to sympathetic vasoconstrictor agents and the subsequent irreversible fall in blood pressure. In addition, sepsis, like other inflammatory conditions, results in a large increase in the production of free radicals, including superoxide anions (O(2)) within the body. Here we show that O(2) reacts with catecholamines deactivating them in vitro. Moreover, this deactivation would appear to account for the hyporeactivity to exogenous catecholamines observed in sepsis, because administration of a superoxide dismutase (SOD) mimetic to a rat model of septic shock to remove excess O(2) restored the vasopressor responses to norepinephrine. This treatment with the SOD mimetic also reversed the hypotension in these animals; suggesting that deactivation of endogenous norepinephrine by O(2) contributes significantly to this aspect of the vascular crisis. Indeed, the plasma concentrations of both norepinephrine and epinephrine in septic rats treated with the SOD mimetic were significantly higher than in untreated rats. Interestingly, the plasma concentrations for norepinephrine and epinephrine were inversely related to the plasma concentrations of adrenochromes, the product of the autoxidation of catecholamines initiated by O(2). We propose, therefore, that the use of a SOD mimetic represents a new paradigm for the treatment of septic shock. By removing O(2), exogenous and endogenous catecholamines are protected from autoxidation. As a result, both hyporeactivity and hypotension are reversed, generation of potentially toxic adrenochromes is reduced, and survival rate is improved.

Molecular aspects of Huntington's disease.

Huntington's disease (HD) is a progressive neurodegenerative disease striking principally medium spiny GABAergic neurons of the caudate nucleus of the basal ganglia. It affects about one in 10,000 individuals and is transmitted in an autosomal dominant fashion. The molecular basis of the disease is expansion of the trinucleotide CAG in the first exon of a gene on chromosome four. The CAG repeats are translated to polyglutamine repeats in the expressed protein, huntingtin. The normal function of huntingtin remains incompletely characterized, but based upon recently defined protein-protein interactions, it appears to be associated with the cytoskeleton and required for neurogenesis. Huntingtin has been demonstrated to interact with such proteins as HAP1, HIP1, microtubules, GADPH, calmodulin, and an ubiquitin-conjugating enzyme. Polyglutamine expansion alters many of these interactions and leads to huntingtin aggregation and the formation of neuronal nuclear inclusions, ultimately culminating in cell death. In this review, we discuss the molecular aspects of HD, including the present understanding of huntingtin-protein interactions, studies with transgenic mice, and postulated mechanisms of huntingtin aggregation.

Neuropeptide Y receptors involved in calcium channel regulation in PC12 cells.

Our laboratory has previously used NGF-differentiated PC12 cells as a sympathetic neuronal model to investigate the effects of NPY on catecholamine synthesis and release. We have additionally used these cells to demonstrate the NPY-induced inhibition of Ca2+ channels which was suggested by those studies. In the present work, multiple NPY, PYY, and PP analogs are utilized to further define the receptor subtypes involved in this Ca2+ channel modulation. We find that in PC12 cells NPY and PP modulate Ca2+ channels through Y1, Y2, Y3, and Y4 receptors. In addition, we show that these receptors are differentially coupled to N, L, and non-N, non-L Ca2+ channel subtypes. The results of the present study in combination with our previous investigations demonstrate an intriguing and complex role for NPY and PP in the modulation of sympathetic neurotransmission.

Neuropeptide Y inhibition of calcium channels in PC-12 pheochromocytoma cells.

We previously demonstrated, using rat PC-12 pheochromocytoma cells differentiated to a sympathetic neuronal phenotype with nerve growth factor (NGF), that neuropeptide Y (NPY) inhibits catecholamine synthesis as well as release. Inquiry into the mechanisms of these inhibitions implicated distinct pathways involving reduction of Ca2+ influx through voltage-activated Ca2+ channels. In the present investigation the effects of NPY on whole cell Ba2+ currents were examined to obtain direct evidence supporting the mechanisms suggested by those studies. NPY was found to inhibit the voltage-activated Ba2+ current in NGF-differentiated PC-12 cells in a reversible fashion with an EC50 of 13 nM. This inhibition was pertussis toxin sensitive and resulted from NPY modulation of L- and N-type Ca2+ channels. The inhibition of L-type channels was not seen with < 1 nM free intracellular Ca2+ or when protein kinase C (PKC) was inhibited by chelerythrine or PKC-(19-31). Furthermore, the effect of NPY on L-type channels was mimicked by the PKC activator phorbol 12-myristate 13-acetate. These studies demonstrate that, in addition to inhibition of N-type Ca2+ channels, in NGF-differentiated PC-12 cells NPY inhibits L-type Ca2+ channels via an intracellular Ca(2+)- and PKC-dependent pathway.

Neuropeptide Y gene expression in immortalized rat hippocampal and pheochromocytoma-12 cell lines.

Employing clonal cell lines derived from rat embryonic hippocampal cells, we detected neuropeptide Y (NPY) mRNA in three progenitor subcloned cell lines. These cell lines upon differentiation express markers indicative of commitment to either neuronal (H19-7; NF +, GFAP -), glial (H19-5; GFAP +, NF -), or bipotential (H583-5; NF +, GFAP + ) lineages. Induction of differentiation was associated with the persistence of the NPY mRNA, however, in the differentiated H19-7 cells a 20-fold increase in NPY mRNA levels was observed (P<0.05). NPY immunoreactivity was observed only in cells with a differentiated neuronal phenotype. The cellular radioimmunoassayable NPY peptide levels increased twelve-fold without a change in extracellular NPY peptide levels by multi-factorially induced neuronal or glial cell differentiation. The differentiated H19-5 cells expressed lower levels of NPY that could not be immunocytochemically detected. The peripheral sympathetic PC-12 neuronal cells examined in the undifferentiated and nerve growth factor-driven differentiated states expressed NPY only upon differentiation. We conclude that NPY is expressed by the cultured undifferentiated and differentiated rat hippocampal clonal cell lines, while the peripheral sympathetic PC-12 neuronal cell line only expresses the NPY gene upon differentiation. These immortalized embryonic neural cell line(s) will provide a hippocampal cell line(s) to conduct future in-vitro investigations targeted at determining the cellular and molecular mechanisms governing NPY gene expression.

Autoreceptor-induced inhibition of neuropeptide Y release from PC-12 cells is mediated by Y2 receptors.

Pheochromocytoma (PC)-12 cells express Y1, Y2, and Y3 neuropeptide Y (NPY) receptors when differentiated with nerve growth factor (NGF). The present work evaluated NGF-differentiated PC-12 cells as a model system to study modulation of NPY release by NPY autoreceptors. We demonstrated that both K+ and nicotine stimulated concomitant release of NPY and dopamine from differentiated PC-12 cells. We also showed in this study that NPY release from PC-12 cells was attenuated in a concentration-dependent manner by peptide YY (PYY)-(13-36), a selective agonist for the Y2 type of NPY receptors. This result demonstrated that NPY release could be modulated by NPY autoreceptors of the Y2 subtype. The inhibitory action of PYY-(13-36) may be mediated at least in part by inhibition of N-type Ca2+ channels, because PYY-(13-36) could not produce further inhibitory effects in the presence of a maximum effective concentration of omega-conotoxin, an N-type Ca2+-channel blocker. The inhibition by PYY-(13-36) could be blocked by pretreatment of cells with pertussis toxin, suggesting that an inhibitory GTP-binding protein was involved. Furthermore, the function of NPY autoreceptors could be modulated by other receptors such as beta-adrenergic and ATP receptors. The evoked release of NPY was also attenuated by ATP and adenosine, which have been shown to be colocalized and coreleased with NPY from sympathetic nerve terminals. These results suggest that PC-12 cells differentiated with NGF may be an ideal model to study regulatory mechanisms of NPY release and that autoreceptor-mediated regulation of NPY release appears to act through the Y2 subtype of the NPY receptor.

Maternal diabetes-induced hyperglycemia and acute intracerebral hyperinsulinism suppress fetal brain neuropeptide Y concentrations.

We examined the effect of streptozotocin-induced maternal diabetes of 6-day duration and 4- to 24-h intracerebroventricular and systemic hyperinsulinism on fetal brain neuropeptide Y (NPY) synthesis and concentrations. Maternal diabetes (n = 6) leading to fetal hyperglycemia (5-fold increase; P < 0.05) and normoinsulinemia caused a 40% decline (P < 0.05) in fetal brain NPY messenger RNA (mRNA) and a 50% decline (P < 0.05) in NPY radioimmunoassayable levels compared to levels in streptozotocin-treated nondiabetic (n = 7) and vehicle-treated control (n = 8) animals. In contrast, systemic hyperinsulinemia (n = 7) of 5- to 100-fold increase (P < 0.05) over the respective control (n = 7) with normoglycemia caused an insignificant (20-30%) decrease in fetal brain NPY mRNA and protein concentrations. However, fetal intracerebroventricular hyperinsulinism (n = 7) with no change in fetal glucose concentrations caused a 50-60% decline (P < 0.05) in only the NPY peptide levels, with no change in the corresponding mRNA amounts. We conclude that fetal hyperglycemia of 6-day duration and intracerebroventricular hyperinsulinism of 4-24 h suppress fetal brain NPY concentrations, the former by a pretranslational and the latter by either a translational/posttranslational mechanism or depletion of intracellular secretory stores. We speculate that fetal hyperglycemia and intracerebroventricular hyperinsulinism additively can inhibit various intrauterine and immediate postnatal NPY-mediated biological functions.

Neuropeptide Y-ATP interactions and release at the vascular neuroeffector junction.

1. The ability of neuropeptide Y (NPY) to potentiate the contractile effect of ATP was examined using the perfused mesenteric arterial bed as a model of the vascular neuroeffector junction. 2. NPY (10(-9)-10(-7) M) and the NPY-Y1 selective agonist, Leu31Pro34 NPY (10(-9)-10(-7) M) both produced a concentration dependent potentiation of the ATP (1 and 3 mM) induced increase in perfusion pressure while the NPY-Y2 selective agonist, NPY 14-36 did not. 3. The NPY-Y1 selective antagonist BIBP 3226 (10-100 nM) produced a significant concentration dependent blockade of the Leu31Pro34 NPY (30 nM) induced potentiation of the ATP (3 mM) induced increase in perfusion pressure. These results are consistent with the NPY-induced potentiation of ATP effect being due to activation of the NPY-Y1 receptor subtype. 4. Periarterial nerve stimulation (supramaximal voltage, 8 and 16 Hz, 30s caused a release of ATP, as well as metabolites, from the perfused mesenteric arterial bed. KCl evoked (50 mM, 5 min) release of ATP from nerve growth factor (NGF) differentiated PC12 cells. 5. Endothelin-1 (ET-1) produced a concentration dependent (10(-15)-10(-8) M) inhibition of the K-1-evoked release of ATP from NGF-differentiated PC12 cells. This effect was mimicked by the selective ETB agonists, BQ 3020, STX-6C and IRL 1620. The ETA/ETB antagonist PD142893 blocked the inhibitory effect of ET-1. These results are consistent with the ET-1 induced inhibition of the evoked release of ATP being due to activation of ETB receptors.

Mechanism of catecholamine synthesis inhibition by neuropeptide Y: role of Ca2+ channels and protein kinases.

We have previously demonstrated that neuropeptide Y (NPY) inhibits depolarization-stimulated catecholamine synthesis in rat pheochromocytoma (PC12) cells differentiated to a sympathetic neuronal phenotype with nerve growth factor (NGF). The present study uses multiple selective Ca2+ channel and protein kinase agonists and antagonists to elucidate the mechanisms by which NPY modulates catecholamine synthesis as determined by in situ measurement of DOPA production in the presence of the decarboxylase inhibitor m-hydroxybenzylhydrazine (NSD-1015). The L-type Ca2+ channel blocker nifedipine inhibited the depolarization-induced stimulation of DOPA production by approximately 90% and attenuated the inhibitory effect of NPY. In contrast, the N-type Ca2+ channel blocker omega-conotoxin GVIA inhibited neither the stimulation of DOPA production nor the effect of NPY. Antagonism of Ca2+/calmodulin-dependent protein kinase (CaM kinase) greatly inhibited the stimulation of DOPA production by depolarization and prevented the inhibitory effect of NPY, whereas alterations in the cyclic AMP-dependent protein kinase pathway modulated DOPA production but did not prevent the effect of NPY. Stimulation of Ca2+/phospholipid-dependent protein kinase (PKC) with phorbol 12-myristate 13-acetate (PMA) did not affect the basal rate of DOPA production in NGF-differentiated PC12 cells but did produce a concentration-dependent inhibition of depolarization-stimulated DOPA production. In addition, NPY did not produce further inhibition of DOPA production in the presence of PMA, and the inhibition by both PMA and NPY was attenuated by the specific PKC inhibitor chelerythrine. These results indicate that NPY inhibits Ca2+ influx through L-type voltage-gated Ca2+ channels, possibly through a PKC-mediated pathway, resulting in attenuation of the activation of CaM kinase and inhibition of depolarization-stimulated catecholamine synthesis.

Amylin-induced relaxation of the perfused mesenteric arterial bed: meditation by calcitonin gene-related peptide receptors.

Amylin is a 37-amino acid peptide that shares considerable homology with calcitonin gene-related peptide (CGRP). Both peptides exert glycoregulatory actions and produce vasodilation of the cardiovascular system. We wished to determine if amylin exerts vasodilatory action in the perfused mesenteric arterial bed in a manner similar to that of CGRP and if so, to determine if amylin and CGRP share a common mechanism of action. Amylin 10(-8), 10(-7), and 10(-6)M produced significant decreases in perfusion pressure by 18, 34, and 45 mm Hg, respectively, of the perfused mesenteric arterial bed pretreated with guanethidine (7 x 10(-6)M) and precontracted with methoxamine (10(-6)-10(-5)M). Amylin was approximately 10 times less potent than CGRP. This vasodilatory effect was not antagonized by atropine in a concentration (10(-6)M) that blocked the vasodilatory action of acetylcholine (ACh) or of nadolol in a concentration that blocked the response to isoproterenol (ISO 10(-6)M). In contrast, the CGRP receptor antagonist [8-37]hCGRP blocked the response of both amylin and CGRP while failing to block the effect of ISO. The depressor effects of CGRP (10(-8)M), amylin (10(-6)M), and ISO (10(-5)M) were 38, 43, and 42 mm Hg without and 5, 12, and 44 mm Hg with [8-37]hCGRP (10(-7)M), respectively. Simultaneous administration of CGRP and amylin failed to produce an additive effect. The depressor effects of CGRP (10(-8)M), amylin (10(-7)M), and CGRP (10(-8)M) plus amylin (10(-7)M) were 50, 32, and 45 mm Hg, respectively. We conclude that amylin exerts a vasodilator action in the perfused mesenteric arterial bed by acting on CGRP1 receptors and suggest that this glycoregulatory hormone may also exert regulatory actions in the vasculature in a manner similar to that of CGRP.

Neuropeptide Y inhibits depolarization-stimulated catecholamine synthesis in rat pheochromocytoma cells.

In PC12 rat pheochromocytoma cells differentiated with nerve growth factor (NGF), neuropeptide Y inhibited depolarization-stimulated catecholamine synthesis as determined by in situ measurement of 3,4-dihydroxyphenylalanine (DOPA) production in the presence of the decarboxylase inhibitor m-hydroxybenzylhydrazine (NSD-1015). The inhibition by neuropeptide Y was concentration-dependent and was prevented by pretreatment with pertussis toxin, suggesting the involvement of a GTP-binding protein of the Gi or Go subtype. The neuropeptide Y analog [Leu31,Pro34]neuropeptide Y also caused inhibition of DOPA production, but was less potent than neuropeptide Y itself, while peptide YY and neuropeptide Y-(13-36) had no significant effect. This pattern is most consistent with the involvement of the neuropeptide Y Y3 receptor subtype. In PC12 cells differentiated with dexamethasone, neuropeptide Y also caused a concentration-dependent inhibition of DOPA production, while peptide YY was again without effect. Neuropeptide Y had no effect on DOPA production in undifferentiated PC12 cells. These results indicate that neuropeptide Y can modulate catecholamine synthesis in addition to its modulatory effects on catecholamine release.

Neuropeptide-Y-ATP interactions at the vascular sympathetic neuroeffector junction.

Neuropeptide Y (NPY) and ATP are considered cotransmitters with norepinephrine (NE) in sympathetic neurons innervating some blood vessels, including those of the mesentery. A prominent action of NPY is to potentiate the postjunctional contractile effect of NE as well as that of other vasoactive agents. We wished to investigate whether NPY also potentiates the contractile effect of ATP and, if so, to determine which receptor subtype mediates such an effect. The effect of NPY, the NPY-Y1-selective agent Leu31Pro34 NPY, and the NPY-Y2-selective fragment NPY 14-36 on the increase in perfusion pressure produced by ATP was examined in rat perfused mesenteric arterial bed. Results demonstrated that both NPY and Leu31Pro34 NPY but not NPY 14-36 potentiated the increase in perfusion pressure produced by ATP. These results suggest that NPY acts on Y1 receptors to enhance the postjunctional response of ATP. The putative NPY antagonist PYX2, but not the putative antagonists benextramine or PYX1, attenuated the effect of NPY, indicating that PYX2 acts as an NPY antagonist in this system. A major action of NPY is to enhance the postjunctional response of both cotransmitters, ATP and NE at the vascular sympathetic neuroeffector junction in the mesenteric arterial bed, and this may be mediated by NPY-Y1 receptors.

A new perspective on the inhibitory role of nitric oxide in sympathetic neurotransmission.

By using, as a model of sympathetic neurons, immortalized rat pheochromocytoma (PC12) cells differentiated by nerve growth factor (NGF), the effect of nitric oxide on sympathetic neurotransmission was examined. The NO donor sodium nitroprusside (SNP; 10(-4)-3 x 10(-4) M) caused an apparent inhibition of dopamine release from PC12 cells, as measured by HPLC. Studies, in the absence of cells, involving the incubation of dopamine (20 ng/ml) or norepinephrine (15 ng/ml) with SNP (10(-6)-3 x 10(-4) M) or authentic NO (6 x 10(-6)-3 x 10(-5) M) revealed a similar reduction in the detection of the catecholamines. In addition, absorption spectroscopy studies showed dopamine and norepinephrine to be oxidized by NO resulting in the formation of their respective quinone products. These observations, coupled with the finding that the ability of dopamine to raise cAMP levels within PC12 cells was reduced after incubation with SNP, reveal that NO inhibits the biological activity rather than the release of catecholamines.

Overflow of endogenous norepinephrine from PVH nucleus of DOCA-salt hypertensive rats.

Previous studies from this laboratory demonstrated that there was enhanced basal and evoked (K+ depolarization) overflow of endogenous norepinephrine (NE) into the perfusate of a push-pull cannula placed in the paraventricular nucleus of the hypothalamus (PVH) of conscious freely moving spontaneously hypertensive rat (SHR) compared with Wistar-Kyoto (WKY) or Sprague-Dawley (SD) rats. The present study was carried out to determine whether results obtained with SHR were specific to this genetic model of hypertension by examining NE release in deoxycorticosterone acetate (DOCA)-salt hypertension. DOCA-salt hypertension was produced in 8-wk-old uninephrectomized SD rats by administering a 50-mg DOCA Silastic pellet subcutaneously 7 days postnephrectomy and providing 0.9% NaCl + 0.2% KCl drinking solution at libitum for 3 wk. Sham-implanted animals received normal tap water. Blood pressure was similar to that of 8- to 10-wk-old SHR. Basal release of NE as well as release after K+ added to the push-pull cannula or sodium nitroprusside or phenylphrine administered intravenously was determined. It was observed that there was no difference in basal overflow or after K+ administration in DOCA-salt hypertensive rats compared with sham animals. Similarly, the increase in NE overflow due to sodium nitroprusside or the decrease due to phenylphrine was similar between DOCA-salt rats or sham controls. This was in sharp contrast to what was observed in SHR: basal or K(+)-evoked release was significantly greater in SHR than WKY, SD, DOCA-salt, or DOCA-sham controls. It is concluded that central noradrenergic activity involving the PVH is not altered in DOCA-salt hypertension.(ABSTRACT TRUNCATED AT 250 WORDS)