Expression and expressional control of nitric oxide synthases in various cell types.
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Animals↗
Biomedical subjects
N J Dun
Publications and source records attributed to N J Dun.
Calcium-activated release of nitric oxide potentiates excitatory synaptic potentials in immature rat sympathetic preganglionic neurons.
1. Whole cell patch recordings were made from sympathetic preganglionic neurons (SPNs), the majority of which contain brain nitric oxide synthase (bNOS), in transverse spinal cord slices of 12- to 16-day-old rats. 2. Repetitive discharge of SPNs induced by a train of depolarizing current pulses (40 Hz. 10 s) was followed by a long-lasting increase (140 +/- 22%, mean +/- SD) of the amplitude of excitatory postsynaptic potentials (EPSPs) evoked by stimulation of lateral funiculus in 50 of 75 SPNs. 3. In slices pretreated with the nitric oxide synthase inhibitors, NG-monomethyl-L-arginine (L-NMA: 100 microM) or Nw-nitro-L-arginine (L-NARG; 30 microM) or with bovine hemoglobin (100 microM), repetitive discharge of SPNs was not followed by a significant increase of EPSPs. 4. Superfusing the slices with L-arginine (L-Arg, 300 microM) but not D-Arg reversibly increased the EPSPs by an average of 140 +/- 19%. 5. Inclusion of the Ca2+ chelator 1.2-bis(2-aminophenoxy)- ethane-N,N,N',N'-tetraacetic acid (BAPTA, 1 mM) in the patch electrodes resulted in no significant increase of EPSPs after repetitive discharge in all cells studied. 6. It is concluded that during repetitive discharge of SPNs, Ca2+ influx via voltage-gated channels activates bNOS, resulting in a release of nitric oxide and potentiation of EPSPs.
Adrenergic Fibers↗
Endogenous and exogenous nitric oxide inhibits norepinephrine release from rat heart sympathetic nerves.
This study was designed to elucidate whether nitric oxide (NO) controls norepinephrine (NE) release from sympathetic nerves of the rat heart. Hearts were perfused in the Langendorff mode with Tyrode's solution. The right sympathetic nerve was stimulated with trains of 1 or 3 Hz and NE release was measured. The NO synthase (NOS) inhibitor NG-nitro-L-arginine (L-NNA) enhanced the evoked NE release in a concentration-dependent manner. This facilitation was independent of the increase in perfusion pressure and was stereospecifically reversed by L-arginine but not D-arginine. Another NOS inhibitor, NG-methyl-L-arginine, produced a similar increase in NE release. The NO-donor compound S-nitroso-N-acetyl-D,L-penicillamine, added in the presence of L-NNA, restored the suppression of NE release in a concentration-dependent fashion. A similar suppression was achieved with 3-morpholinosydnonimine. These results demonstrated that NE release is under the inhibitory control of endogenous NO. Western blots demonstrated the presence of neuronal NOS I and endothelial NOS III in the hearts. Perfusion of the hearts with a low concentration of the detergent CHAPS produced functional damage of the endothelium, as evidenced by an increase in perfusion pressure and a conversion of the acetylcholine-induced coronary vasodilation to a constriction. However, CHAPS treatment did not produce a facilitation of NE release (as did the NOS inhibitors), and L-NNA still increased NE release in CHAPS-treated hearts. Double-labeling immunofluorescence histochemistry showed NOS I immunoreactivity in stellate ganglion cells and in neurons of the heart, some of which also stained positive for tyrosine hydroxylase.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Infrequent co-localization of nitric oxide synthase and calcium binding proteins immunoreactivity in rat neocortical neurons.
Immunoreactivity to nitric oxide synthase (NOS-IR) was detected in a small population of rat neocortical neurons scattered throughout the frontal, parietal, temporal and occipital cortices. Two-color double-labeling studies revealed that the number of NOS-IR neocortical neurons expressing immunoreactivity to the calcium binding proteins parvalbumin (PV-IR), calbindin-D28K (CB-IR) or calretinin (CR-IR), was low. The absence or low level of calcium binding proteins may imply a reduced Ca2+ buffering capacity in NOS-containing neurons, thereby contributing to their vulnerability to Ca2+ influx through the activated AMPA/kainate receptors reported by others.
Amino Acid Oxidoreductases↗
c-fos antisense attenuates Fos expression in rat central neurons induced by hemorrhage.
Hemorrhage caused by withdrawing 3-4 cm3 of blood from adult rats over a period of 1-2 h induced the expression of Fos proteins, which were detected by immunocytochemical methods in neurons of the hypothalamic supraoptic (SON) and paraventricular (PVN) nuclei, area postrema, nucleus of the solitary tract (NTS), ventrolateral medulla (VLM) and in intermediolateral cell column (IML) of the spinal cord. Daily intraventricular (i.c.v.) injections of c-fos antisense (50 nmol 5 microliters -1) for 2 days prior to hemorrhage markedly attenuated the extent and intensity of Fos-immunoreactivity (FOS-IR) compared with that of rats injected with c-fos sense in the areas mentioned above. The results indicate the effectiveness of i.c.v. administration of c-fos antisense in blocking Fos expression in central neurons following hemorrhage.
Animals↗
Serotonergic suppression of interhemispheric cortical synaptic potentials.
The inhibitory effects of 5-hydroxytryptamine (5-HT) on interhemispheric and intracortical synaptic potentials in layer V neurons of the rat medial prefrontal (MFC) cortex were examined. Low concentrations (1-3 microM) of 5-HT selectively attenuated polysynaptic potentials that were similarly evoked by callosal or white matter stimulation. Maximally effective concentrations of 5-HT blocked interhemispheric transmission by 50-90%, as evidenced by an attention of the short latency callosal depolarizing synaptic potential (e-DPSP). These effects of 5-HT were not associated with a change in membrane potential or input resistance. The e-DPSP was characterized as having an N-methyl-D-aspartate (NMDA) and a non-NMDA component; the non-NMDA component was attenuated by 5-HT. Attenuation of the synaptic potential was accompanied by an attenuation of a postsynaptic glutamate potential. Suppression of both the e-DPSP and the glutamate potential was concentration dependent with 10-100 microM being maximally effective. The 5-HT1A/2 antagonist, spiperone, antagonized the effects of 5-HT on synaptic and glutamate potentials. Spiperone (1 microM) shifted the concentration-effect curves for suppression of the e-DPSP and the glutamate potential to the right; however, the Kb for the glutamate potential concentration-effect curve was 10 times that for the e-DPSP curve. The differential antagonist sensitivity of synaptic and glutamate potentials was an indication that serotonin acted on more than one receptor subtype to reduce interhemispheric transmission.
2-Amino-5-phosphonovalerate↗
Colocalization of nitric oxide synthase and somatostatin immunoreactivity in rat dentate hilar neurons.
Distribution of nitric oxide synthase (NOS), somatostatin (SSN), and parvalbumin (PV) was studied in the rat hippocampus by immunohistochemical methods. The aim was to explore the interrelationship between SSN-immunoreactive (SSN-IR) neurons in the dentate hilus, which have been shown to be vulnerable to a number of pathophysiological insults, and the presence or absence of NOS and/or PV in the same subset of dentate hilar neurons. Small NOS-IR neurons were scattered in the pyramidal, oriens, and radiatum layers of the CA1-CA3 areas and in the subiculum, where larger NOS-IR neurons were occasionally noted. In the area dentata, NOS-IR neurons, which were composed of small and large polymorphic cells, appeared as a single file at the hilar border with the granule cell layer and clustered in the hilus in fairly high density. Double-labeling techniques showed that most NOS-IR neurons in the hilus were SSN-IR, whereas coexistence of NOS and PV immunoreactivity or SSN and PV immunoreactivity was low in dentate hilar neurons. In other areas of the hippocampus, colocalization of NOS and SSN in the same neurons was much less frequent. Thus, SSN-IR neurons in the dentate hilus constitute a population of neurons that contain the enzyme NOS as well. The presence of NOS coupled to the lack or low level of PV in this group of neurons may provide a neurochemical basis for their high susceptibility to certain pathophysiological insults.
Amino Acid Oxidoreductases↗
Nitric oxide synthase immunoreactivity in rat pontine medullary neurons.
Nitric oxide synthase immunoreactivity was detected in neurons and fibers of the rat pontine medulla. In the medulla, nitric oxide synthase-positive neurons and processes were observed in the gracile nucleus, spinal trigeminal nucleus, nucleus of the solitary tract, dorsal motor nucleus of the vagus, nucleus ambiguus, medial longitudinal fasciculus, reticular nuclei and lateral to the pyramidal tract. In the pons, intensely labeled neurons were observed in the pedunculopontine tegmental nucleus, paralemniscal nucleus, ventral tegmental nucleus, laterodorsal tegmental nucleus, and lateral and medial parabrachial nuclei. Labeled neurons and fibers were seen in the interpeduncular nuclei, dorsal and median raphe nuclei, central gray and dorsal central gray, and superior and inferior colliculi. Double-labeling techniques showed that a small population (< 5%) of nitric oxide synthase-positive neurons in the medulla also contained immunoreactivity to the aminergic neuron marker tyrosine hydroxylase. The majority of nitric oxide synthase-immunoreactive neurons in the dorsal and median raphe nuclei were 5-hydroxytryptamine-positive, whereas very few 5-hydroxytryptamine-positive cells in the caudal raphe nuclei were nitric oxide synthase-positive. Virtually all nitric oxide synthase-positive neurons in the pedunculopontine and laterodorsal tegmental nuclei were also choline acetyltransferase-positive, whereas nitric oxide synthase immunoreactivity was either low or not detected in choline acetyltransferase-positive neurons in the medulla. The results indicate a rostrocaudal gradient in the intensity of nitric oxide synthase immunoreactivity, i.e. it is highest in neurons of the tegmentum nuclei and neurons in the medulla are less intensely labeled. The majority of cholinergic and serotonergic neurons in the pons are nitric oxide synthase-positive, whereas the immunoreactivity was either too low to be detected or absent in the large majority of serotonergic, aminergic and cholinergic neurons in the medulla.
Amino Acid Oxidoreductases↗
Spontaneous and transmitter-induced rhythmic activity in neonatal rat sympathetic preganglionic neurons in vitro.
1. Intracellular recordings were made from sympathetic preganglionic neurons (SPNs) in transverse thoracolumbar spinal cord slices of 12- to 16-day-old rats. 2. A population of SPNs exhibited spontaneous, rhythmic activities that can be grouped into regular firing, bursting, or tonic oscillatory mode; the spikes were interspersed with subthreshold afterdepolarizing potentials (ADPs). 3. Spontaneous activities were not abolished by the excitatory amino acid antagonists 6-cyano-7-nitroquinoxoline-2,3-dione (CNQX, 10-20 microM) and D-2-amino-5-phosphonovaleric acid (APV, 10-20 microM) nor by the inhibitory amino acid antagonists bicuculline (10-30 microM) and strychnine (1 microM). 4. Stimulation of lateral funiculus elicited a CNQX-sensitive excitatory postsynaptic potential (EPSP) in the majority of spontaneously active SPNs tested. Moreover, rhythmic activities could be triggered by stimulation of lateral funiculus in 8 of the 80 otherwise quiescent SPNs tested. 5. Spontaneous spikes and ADPs were abolished by either Na(+)-free or tetrodotoxin (TTX, 0.5 microM)-containing Krebs solution. 6. The spontaneous spike afterhyperpolarization (AH) consisted of two components: a fast AHf and a slower AHS with a mean half decay time of 74.5 +/- 10.2 and 153.8 +/- 15.4 (SD) ms, respectively. 7. Superfusing the slices with a Ca(2+)-free Krebs solution or Krebs solution containing tetraethylammonium (TEA, 10-30 mM) preferentially blocked the AHS, thereby increasing the spontaneous discharge frequency. 8. Caffeine (3 mM) initially prolonged the AHS, thereby slowing the discharge frequency; the spontaneous discharges were eventually obliterated in the continuous presence of caffeine.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Nitric oxide synthase immunoreactivity in rat superior cervical ganglia and adrenal glands.
Nitric oxide synthase-immunoreactivity (NOS-IR) was detected in strands of nerve fibers entering the rat superior cervical ganglia (SCG) and in nerve fibers forming a plexus beneath the capsule of adrenal glands. Within the SCG, varicose NOS-IR fibers encircled virtually all postganglionic neurons and small diameter cells, presumably small intensely fluorescent (SIF) cells. Perikarya of SIF cells exhibited strong NOS-IR, whereas the level appeared to be low in postganglionic neurons. Decentralization of the SCG for 4-6 days markedly reduced the number as well as the intensity of NOS-IR fibers without causing a detectable change of NOS-IR in the postganglionic neurons and SIF cells. Beneath the adrenal capsule, bundles of NOS-IR fibers bifurcated and made a sharp turn to reach the adrenal medulla. Chromaffin cells, which themselves exhibited fairly strong NOS-IR, appeared to be surrounded by NOS-IR fibers. The result shows that NOS-IR is present in pre- and post-synaptic elements of the sympathetic ganglia and adrenal medulla, representing a complex system that may regulate the activity of ganglionic neurons and chromaffin cells via a number of sites of action.
Adrenal Glands↗
Hemorrhage induces Fos immunoreactivity in rat medullary catecholaminergic neurons.
In urethane anesthetized rats one hour after lowering the systolic blood pressure to 70-75 mmHg by withdrawing 3-4 ml of blood, Fos immunoreactivity (Fos-IR), confined to the cell nucleus, was detected bilaterally in numerous cells of the nucleus of the solitary tract (NTS) and ventrolateral medulla (VLM). A few Fos-IR neurons were observed in the lateral reticular nucleus, dorsal medullary reticular nucleus, spinal trigeminal nucleus, medial inferior olive, interfasciculus hypoglossi and paramedian rostral medulla. In sham-operated animals, a much smaller number of Fos-IR neurons were scattered in the NTS, VLM and other nuclei mentioned above. Double labeling with antisera to tyrosine-hydroxylase (TH) and phenylethanolamine-N-methyltransferase (PNMT) showed that 60% of TH-positive neurons in the NTS contained Fos-IR, and 70-80% of TH-positive neurons in the caudal VLM and 50-60% of PMNT-positive neurons in the rostral VLM expressed Fos-IR. Only a few TH- or PNMT-positive neurons in the C2, C3 (paramedian rostral medulla) areas and within the medial longitudinal fasciculus were Fos-IR. About 40% of PNMT/Fos-IR neurons in the rostral VLM contained the retrograde tracer fluorogold, which was injected (< 1 microliter) into the white matter dorsolateral to the intermediolateral cell column of T2-T3 segments 2 to 3 days prior to hemorrhagic experiments. Very few TH-positive neurons in the caudal VLM contained fluorogold. Finally, clusters of Fos-IR neurons, which also labeled with antisera to choline acetyltransferase, were detected in the intermediolateral cell column of the spinal cord. The results indicate that during hemorrhage aminergic neurons in the caudal and rostral VLM and in the NTS are activated insofar as c-fos expression is concerned. As a corollary, the monoaminergic neurons in the medulla constitute an essential component in the ascending as well as descending reflex pathway involved in the adjustment of cardiovascular dynamics during hemorrhage.
Animals↗
Nitric oxide synthase immunoreactivity in the rat, mouse, cat and squirrel monkey spinal cord.
The distribution of nitric oxide synthase-immunoreactive neurons was examined in the spinal cord of rats, mice, cats and squirrel monkeys at the light microscopic level. Some sections were processed for choline acetyltransferase immunoreactivity. Double-labeling techniques were used to assess possible co-localization of nitric oxide synthase and choline acetyltransferase immunoreactivity in the same spinal neurons. Nitric oxide synthase-immunoreactive neurons were concentrated in three fairly well-defined regions of the spinal cord of all species studied: (i) the intermediolateral cell column of the thoracic and sacral segments, (ii) lamina X of all segments, and (iii) the superficial layers of the dorsal horn of all segments. A few nitric oxide synthase-immunoreactive neurons were scattered in the deeper laminae and the ventral horn. There were fewer nitric oxide synthase-positive neurons in monkey spinal lamina X and dorsal horn than in similar locations of rodents and felines. Double-staining showed that not all choline acetyltransferase-positive neurons in the intermediate cell column and lamina X were nitric oxide synthase-immunoreactive. In the ventral horn, choline acetyltransferase-positive neurons (presumed motoneurons) were nitric oxide synthase-negative. In addition to cell bodies, nitric oxide synthase-positive fibers were scattered in the dorsal, lateral and ventral horns of all species. Finally, punctate nitric oxide synthase-immunoreactive fibers were seen traversing the dorsal, lateral and ventral white matter, and reaching the respective gray matter. The present study shows that, in spite of quantitative differences, the pattern of distribution of nitric oxide synthase-positive neurons in the spinal cord was similar across the four species. The concentration of nitric oxide synthase-positive neurons in the autonomic nuclei and laminae I, II and X of all four species underscores a prominent role of these neurons in visceral and sensory functions.
Amino Acid Oxidoreductases↗
Excitatory amino acids depress synaptic currents in neonate rat sympathetic preganglionic neurons.
1. Whole-cell patch-clamp recordings were made from sympathetic preganglionic neurons (SPNs) in transverse thoracolumbar spinal cord slices of 10- to 16-day-old rats, and the effects of L-glutamate (L-Glu) and analogues on excitatory (EPSCs) and inhibitory (IPSCs) postsynaptic currents evoked by stimulation of lateral funiculus were studied. 2. L-Glu (10-300 microM), quisqualate (QA, 0.1-3 microM), kainate (KA, 0.3-10 microM), ibotenate (10-25 microM), and L-2-amino-4-phosphonobutyrate (L-AP4, 25-300 microM) depressed the EPSCs and IPSCs in a concentration-dependent manner, the rank order being QA > KA > ibotenate > L-AP4 > or = L-Glu. The metabotropic glutamate receptor agonist trans-1-amino-1,3-cyclopentane-dicarboxylic acid (trans-ACPD, 25-100 microM) reduced the synaptic currents as well. A similar effect was not observed with N-methyl-D-aspartate (NMDA). 3. The excitatory amino acid uptake inhibitor L-aspartic acid-beta-hydroxamate (AAH, 100 microM), although having little or no direct effect on EPSCs, unmasked the inhibitory effect of low (< or = 1 microM) concentrations of L-Glu. 4. The synaptic depression was not accompanied by a detectable change in holding currents or EPSC reversal potentials and decay constants in the majority of SPNs studied. At higher concentrations, L-Glu and analogues, but not L-AP4, induced an inward current in some SPNs. 5. Although strongly depressing the EPSCs, L-AP4 and trans-ACPD had no significant effect on the amplitude of inward current induced by exogenous L-Glu.(ABSTRACT TRUNCATED AT 250 WORDS)
Amino Acids↗
Putative transmitter systems of mammalian sympathetic preganglionic neurons.
The sympathetic nervous system evokes complex effects at multiple target organs in response to external, internal as well as mental stimuli. This output involves an interplay between the actions of a number of transmitters and modulators and at the postsynaptic and presynaptic sites of the autonomic ganglia and the sympathetic preganglionic neurons (SPNs). This review concerns particularly the SPNs of the cat and neonatal rat, studied by means of electrophysiological and immunohistochemical methods. Four types of responses may be elicited, the fast EPSP and IPSP, and their currents, and the slow EPSPs and IPSPs, and their currents. Glutamate and glycine appear to mediate the fast excitatory and inhibitory responses, respectively; peptides and amines seem to be responsible for generating the slow excitatory response, while the slow inhibitory response, found so far only in the cat, appears to be mediated by norepinephrine. Finally, glutamate, enkephalin and GABA, but not glycine attenuate the release of the inhibitory and excitatory transmitters from the nerve terminals abutting on the SPNs. The supraspinal efferent and afferent projections which may release the transmitters and modulators in question are discussed, as well the mechanisms that ensure appropriate programming and moment-to-moment regulation of the autonomic output.
Animals↗
Nitric oxide synthase immunoreactivity in rat spinal cord.
Immunoreactivity to nitric oxide synthase (NOS-IR) and choline acetyltransferase (ChAT-IR) was detected in the adult rat spinal cord using the avidin-biotin-peroxidase technique. Intensely stained NOS-positive neurons with cell processes were observed in the intermediolateral cell column of the thoracic and sacral segments and around the central canal of all segments. These areas also contained ChAT-IR neurons. A number of small- to medium-sized NOS-IR cells were noted in the superficial and deeper laminae throughout the entire cord. NOS-IR was not detected in the ventral horn motoneurons, which were, however, ChAT-IR. The results indicate that NOS-IR is present in autonomic preganglionic neurons and in selected neurons in the dorsal horn and lamina X, but appears to be absent in motoneurons.
Amino Acid Oxidoreductases↗
Hypotension preferentially induces c-fos immunoreactivity in supraoptic vasopressin neurons.
Immunoreactivity to Fos protein (Fos-IR) was detected in rat hypothalamic neurons within 1 h of onset of hemorrhage by withdrawing 4-5 ml of blood, which lowered the arterial blood pressure to 50-70 mm Hg. About 70% of vasopressin (AVP)-containing neurons in the supraoptic nucleus (SON) and 20% in the paraventricular nucleus (PVN) expressed Fos-IR. In contrast, 5% of oxytocin (OXY)-containing neurons in the SON and < 1% in PVN were Fos-IR. Intravenous infusion of the vasodilating agent, nitroprusside, which lowered the blood pressure to levels comparable to that attained by hemorrhage, induced Fos-IR in greater than 65% of AVP-containing neurons in the SON, while relatively few AVP neurons in the PVN were Fos positive. These results suggest that hemorrhage or hypotension preferentially induces c-fos expression in supraoptic AVP-containing neurons.
Animals↗
Presynaptic GABAB receptor activation attenuates synaptic transmission to rat sympathetic preganglionic neurons in vitro.
Intracellular recordings were made from sympathetic preganglionic neurons (SPNs) in transverse neonate rat spinal cord slices. Superfusion of gamma-aminobutyric acid (GABA; 25-100 microM) or (-)-baclofen (1-30 microM) consistently attenuated the excitatory postsynaptic potentials (EPSPs) evoked by stimulation of dorsal rootlets or lateral funiculus, without causing a significant change of the resting membrane potential and input resistance of the SPNs or of the depolarizations induced by pressure applications of glutamate; the IC50 for baclofen was 2.5 microM. When superfused at a higher concentration (greater than or equal to 500 microM) or ejected by pressure GABA caused a bicuculline-sensitive membrane hyperpolarization. The enantiomer (+)-baclofen (10-50 microM) and the GABAA agonist muscimol (1-10 microM) had no significant effect on the EPSPs. The GABAB receptor antagonist 2-hydroxy-saclofen caused a 10 fold rightward shift of the baclofen dose-response curve, whereas the GABAA receptor antagonist bicuculline (10-50 microM) was ineffective. Glycine had no significant effects on the EPSPs in the concentrations (10-100 microM) tested here. The results indicate that of the two putative inhibitory transmitters in the spinal cord GABA but not glycine depresses EPSPs evoked in the rat SPNs by acting on presynaptic GABAB receptors, the activation of which results in a reduction of excitatory transmitter release.
Animals↗
Hypovolemia induces Fos-like immunoreactivity in neurons of the rat supraoptic and paraventricular nuclei.
Immunoreactivities to Fos proteins were detected in numerous neurons in the supraoptic, paraventricular and accessory neurosecretory nuclei 1 h following withdrawal of 4-5 cc of blood from the rat femoral arteries. Few or no positive cells were observed in the same nuclei in sham-operated or control animals. It is concluded that hypovolemia induces c-fos expression in hypothalamic neurons known to be associated with blood volume/pressure regulation.
Animals↗