N-methyl-DL-aspartic Acid Research Articles

Contributions of Second- and Third-Order Retinal Neurons to Cone Electroretinograms After Loss of Rod Function in Rhodopsin P347L Transgenic Rabbits.

To determine the contribution of second- and third-order retinal neurons to the photopic electroretinograms (ERGs) after the degeneration of the rods in rhodopsin P347L transgenic rabbits (Tg). Four wild-type (WT) rabbits and four Tg rabbits were studied at 18 months of age. The photopic ERGs elicited at stimulus onset and offset were analyzed. To block different retinal pathways, 2-amino-4-phosphonobutyric acid (APB), 6-cyano-7-nitroquinoxaline-2, 3 (1H,4H)-dione (CNQX), tetrodotoxin (TTX), and N-methyl-DL-aspartic acid (NMDA) were injected intravitreally. Digital subtraction of the postdrug ERGs from the predrug ERGs was used to determine the contributions of the ON-components blocked by APB, the OFF-components blocked by CNQX, and the third-order neurons blocked by TTX+NMDA. Contribution of the cone photoreceptors to the photopic ERGs in Tg rabbits was approximately 10% of that in WT rabbits. The amplitudes of the positive waves of the ON-components at stimulus onset in Tg rabbits were approximately one-half as large as those in WT. On the other hand, the amplitudes of the positive waves of the OFF-components at stimulus offset in Tg rabbits were approximately 1.4 to 2.3 times larger than those in WT. Transgenic rabbits had a positive wave at stimulus offset, which was reduced after the TTX+NMDA injection. A reduced ON-component and an augmented OFF-component with abnormal responses of the third-order neurons contributed to the cone ERGs after the loss of rod function in Tg rabbits. Our results suggest a complex synaptic remodeling of the residual retinal cells in the advanced stage in Tg rabbits.

Suprachiasmatic Nucleus and Subparaventricular Zone Lesions Disrupt Circadian Rhythmicity but Not Light-Induced Masking Behavior in Nile Grass Rats.

The ventral subparaventricular zone (vSPVZ) receives direct retinal input and influences the daily patterning of activity in rodents, making it a likely candidate for the mediation of acute behavioral responses to light (i.e., masking). We performed chemical lesions aimed at the vSPVZ of diurnal grass rats (Arvicanthis niloticus) using N-methyl-D,L-aspartic acid (NMA), a glutamate agonist. Following NMA lesions, we placed grass rats in various lighting conditions (e.g., 12:12 light-dark, constant dark, constant light); presented a series of light pulses at circadian times (CT) 6, 14, 18, and 22; and placed them in a 7-h ultradian cycle to assess behavioral masking. Extensive bilateral lesions of the vSPVZ disrupted the expression of circadian rhythms of activity and abolished the circadian modulation of masking responses to light, without affecting light-induced masking behavior per se. We also found that in diurnal grass rats, NMA was capable of destroying not only neurons of the vSPVZ but also those of the suprachiasmatic nucleus (SCN), even though excitotoxins have been ineffective at destroying cells within the SCN of nocturnal rodents. The vulnerability of the grass rat's SCN to NMA toxicity raises the possibility of a difference in density of receptors for glutamate between nocturnal and diurnal species. In cases in which damage extended to the SCN, masking responses to light were present and similar to those displayed by animals with damage restricted to the vSPVZ. Thus, extensive bilateral lesions of the SCN and vSPVZ disrupted the expression of circadian rhythms without affecting acute responses to light in a diurnal species. Our present and previous results suggest that retinorecipient brain areas other than the SCN or vSPVZ, such as the intergeniculate leaflet or olivary pretectal nucleus, may be responsible for the mediation of masking responses to light in the diurnal grass rat.

Contribution of N-methyl-dl-aspartic acid (NMDA)-sensitive neurons to generating oscillatory potentials in Royal College of Surgeons rats

We investigated how the N-methyl-DL-aspartic acid (NMDA) receptor contributes to generating oscillatory potentials (OPs) of the electroretinogram (ERG) in the Royal College of Surgeons (RCS) rat. Scotopic ERGs were recorded from dystrophic and wild-type congenic (WT) RCS rats (n=20 of each) at 25, 30, 35, and 40days of age. The stimulus intensity was increased from -2.82 to 0.71 log cd-s/m(2) to obtain intensity-response function. NMDA was injected into the vitreous cavity of the right eyes. The left eyes were injected with saline as controls. The P3 obtained by a-wave fitting was digitally subtracted from the scotopic ERG to isolate the P2. For the OPs, the P2 was digitally filtered between 65 and 500Hz. The amplitudes of OP1, OP2, OP3, and OP4 were then measured and summed and designated as ΣOPs. The implicit times of OP1, OP2, and OP3 were also measured. The frequency spectra of the OPs were analyzed using fast Fourier transform (FFT). The maximum ERG a- and b-waves as well as ΣOPs amplitudes reduced with age in dystrophic rats. Compared with intravitreal saline injection, administration of NMDA decreased ΣOPs amplitudes from 30days of age in dystrophic rats, while it did not attenuate ΣOPs amplitudes in WT rats. The implicit times of the OPs of the maximum ERG were prolonged by NMDA injections in WT and dystrophic rats. NMDA/saline ratios of ΣOPs amplitudes area under the FFT curves were significantly lower in dystrophic rats from 30days of age than that in WT rats. In the early stage of photoreceptor degeneration, intravitreal NMDA injection attenuated OPs amplitudes in dystrophic rats. This indicates that NMDA receptors play a significant role in generating OPs amplitudes with advancing photoreceptor degeneration.

Chiral separation of N‐methyl‐dl‐aspartic acid in rat brain tissue as N‐ethoxycarbonylated (S)‐(+)‐2‐octyl ester derivatives by GC‐MS

A selective and sensitive analytical method was developed for enantiomeric separation and determination of N-methyl-DL-aspartic acid (NMA). The method involved the conversion of each enantiomer into N-ethoxycarbonylated (S)-(+)-2-octyl ester derivative for the direct separation by gas chromatography-mass spectrometry (GC-MS). The diastereomeric derivatives showed characteristic mass spectral properties for analysis by selected ion monitoring mode (SIM) and enabling enantioseparation on an achiral capillary column. Two enantiomers were baseline separated, and the detection limits for N-methyl-L-aspartic acid (NMLA) and N-methyl-D-aspartic acid (NMDA) were 0.07 and 0.03 ng/g, respectively. When applied to rat brain tissues for absolute configuration of NMA, only NMDA was determined, while NMLA was monitored as lower than the limit of detection.

Enhancement of ON-Bipolar Cell Responses of Cone Electroretinograms in Rabbits with the Pro347Leu Rhodopsin Mutation

To determine how the different stages of retinal processing change after photoreceptor degeneration in rabbits carrying the Pro347Leu rhodopsin mutation (Tg rabbits). Cone electroretinograms (ERGs) were elicited by 150-ms duration stimuli from 13 Tg rabbits at 12 and 24 weeks of age. The ERG recordings were made before and after an intravitreal injection of tetrodotoxin citrate (TTX) plus N-methyl-dl-aspartic acid (NMDA), with the addition of 2-amino-4-phosphonobutyric acid (APB) and then cis-2,3-piperidine-dicarboxylic acid (PDA) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Digital subtraction of the ERG after the injection from the ERG before the injection was used to extract the components that were blocked by these drugs. Thirteen age-matched, wild-type (WT) rabbits were studied with the same protocol. In Tg rabbits, the cone ERGs elicited by intermediate intensities had a depolarizing pattern. At 12 weeks of age, the photoreceptor and OFF-bipolar/horizontal cell responses reflected in the ERG in the Tg rabbits did not differ significantly from those in the WT rabbits. The ON-bipolar cells and the third-order neuronal responses recorded after pharmacologic blockade were significantly enhanced in the Tg rabbits compared with those recorded in the WT rabbits. At 24 weeks of age, the ERG waveforms representing the photoreceptors and OFF-bipolar/horizontal cell responses were significantly decreased, but those representing the ON-bipolar cell and third-order neuronal responses were still preserved in the Tg rabbits. A depolarizing pattern of the cone ERG responses was seen in Pro347Leu Tg rabbits. The enhancement or preservation of the ON-bipolar cell response in the ERGs contributed to shaping the waveform in the Tg rabbits. In this model, the functional alterations in the ON-pathway took place before the deterioration of cone photoreceptor function.

Pharmacological dissection of multifocal electroretinograms of rabbits with Pro347Leu rhodopsin mutation

To determine whether photoreceptor degeneration in transgenic (Tg) rabbits carrying the Pro347Leu rhodopsin mutation alters the neural activity of the middle and inner retinal neurons. Multifocal electroretinograms (mfERGs) were recorded from eight 12-week-old Tg rabbits both before and after intravitreal injection of the following: tetrodotoxin citrate (TTX), N-methyl-DL: -aspartic acid (NMDA), 2-amino-4-phosphonobutyric acid (APB), and cis-2,3-piperidine-dicarboxylic acid (PDA). Digital subtraction of the mfERGs recorded after the drugs were administered from those recorded before was used to extract the components that were eliminated by these drugs. Eight agematched, wild-type (WT) rabbits were studied with the same protocol. There was no reduction in the amplitude of the cone photoreceptor response of the mfERGs in Tg rabbits. Both the first positive and the first negative waves of the ON-bipolar cell responses were significantly larger in the Tg than in the WT rabbits. Late negative waves of the ON-bipolar cell response were recorded only in the WT rabbits. The first negative wave of the inner retinal responses was larger in the Tg than in the Wt rabbits. The late positive waves were seen mainly in the WT rabbits. The ON-bipolar cell and inner retinal responses were altered at the early stage of photoreceptor degeneration in Tg rabbits despite the preservation of the cone photoreceptor responses.

Microinjection of NMDA Receptor Agents into the Central Nucleus of the Amygdale Alters Water Intake in Rats

The central nucleus of the amygdala (CeA) is a forebrain structure which is important in regulation of ingestive behavior and there is direct and circumstantial evidence to indicate that some circuits involved with feeding behavior include glutamatergic elements. The present study examined whether administration of NMA (N-Methyl-DL-aspartic acid) or MK801 into the CeA altered water intake under deprivation. Materials and Methods Animals were deprived for 24 hr before tested for water intake. NMDA (N-methyl-D-aspartate) glutamatergic receptor agonist, NMA and its antagonist, MK801 were infused bilaterally, and water intake measured for 1 hr thereafter. Results The intra-CeA injection of NMDA glutamatergic agonist, NMA (0.25, 0.5 and 0.75 µg/rat) increased water intake (P<0.05). However, administration of NMDA glutamatergic antagonist, MK801 (0.25, 0.5 and 1 µg/rat) decreased water intake significantly (P<0.05). Conclusion These data suggest that NMDA receptors in the CeA are responsible for the glutamatergic modulation of water intake in this nucleus.

Anticonvulsant activity of Nylandtia spinosa L. Dumont (Polygalaceae) aqueous and methanol leaf extracts in mice

Aqueous and methanol leaf extracts of Nylandtia spinosa L. Dumont (Polygalaceae) were evaluated for anticonvulsant activity against tonic seizures produced in mice by pentylenetetrazole (PTZ), bicuculline, picrotoxin, and N-methyl-DL-aspartic acid (NMDLA). Aqueous leaf extract of N. spinosa (50-400 mg/kg, i.p.) and methanol extract (50-400 mg/kg, i.p.) significantly attenuated PTZ (95 mg/kg, i.p.)-induced tonic seizures. Doses of 400 mg/kg (i.p.) and 100-400 mg/kg (i.p.) of aqueous extract of N. spinosa significantly delayed the onset of tonic seizures elicited by bicuculline (35 mg/kg, i.p.) and picrotoxin (12 mg/kg, i.p.), respectively. Methanol extract (200-400 mg/kg, i.p.) and (50-400 mg/kg, i.p.) significantly delayed the onset of tonic seizures induced by bicuculline (35 mg/kg, i.p.) and picrotoxin (12 mg/kg, i.p.), respectively, whereas 400 mg/kg (i.p.) significantly reduced the incidence of picrotoxin (12 mg/kg, i.p.)-induced seizures. Both aqueous and methanol leaf extracts of N. spinosa did not affect NMDLA (400 mg/kg, i.p.)-induced tonic seizures. Phenobarbitone (12.5 mg/kg, i.p.) and diazepam (0.5 mg/kg, i.p.) antagonized tonic seizures induced by PTZ (95 mg/kg, i.p.), bicuculline (35 mg/kg, i.p.), and picrotoxin (12 mg/kg, i.p.) but did not affect NMDLA (400 mg/kg, i.p.)-induced seizures. Phenytoin (30 mg/kg, i.p.) did not alter the tonic seizures produced by either PTZ (95 mg/kg, i.p.), bicuculline -2-(35 mg/kg, i.p.), or picrotoxin (12 mg/kg, i.p.). The results obtained indicate that both aqueous and methanol leaf extracts of N. spinosa possess anticonvulsant property, thus justifying the use of the plant by traditional medicine practitioners in the treatment of epilepsy. The relatively high LD(50) of greater than 3600 mg/kg (p.o.) and 1780 mg/kg (i.p.) obtained with the aqueous extract suggest that the plant is relatively safe in mice. The phytochemical analysis carried out showed the presence of tannins, saponins, reducing sugars, alkaloids, flavonoids, triterpene steroids, and cardiac glycosides in the plant material.

Photopic ERG Negative Response from Amacrine Cell Signaling in RCS Rat Retinal Degeneration

The authors investigated photopic electroretinographic changes during degeneration in the Royal College of Surgeons (RCS) and transgenic P23H rhodopsin rat models, including the cellular origins of a large corneal-negative component that persists in the RCS rat. Photopic and scotopic electroretinograms (ERGs) were recorded from dystrophic RCS (RCS-p(+)/Lav) rats (4-18 weeks old) and transgenic rhodopsin Pro23His line 1 (P23H) rats (4-30 weeks old). Age-matched congenic (RCS-rdy(+)p(+)/Lav) and Sprague-Dawley rats were used as controls. N-methyl-DL-aspartic acid (NMA), dopamine, and gamma-aminobutyric acid (GABA) were injected intravitreally, and optic nerve sectioning (ONS) was performed to suppress or remove inner retinal neuron activity. Retinal morphology for cone cell counts and immunohistochemistry for quantification of Kir4.1 channels were performed at various stages of degeneration. As degeneration progressed, the photopic ERG of RCS dystrophic rats was distinctly different from that of P23H rats, primarily because of the growth of a corneal-negative response (RCS-NPR) after the b-wave in RCS rats. This response had a peak time similar to the photopic negative response (PhNR) in controls but with a more gradual recovery phase, and it was not affected by ONS. The PhNR in P23H rats declined linearly with the b-wave. NMA and GABA eliminated the RCS-NPR and uncovered a larger b-wave in RCS rats at late stages of degeneration, but NMA had little effect on the ERG in P23H rats. The NMA-sensitive negative response in RCS rats declined with age more slowly than did the NMA-isolated b-wave. The density of Kir4.1 channels at the endfeet of Müller cells and in the proximal retina increased significantly between 6 to 10 weeks and 14 weeks of age in the RCS rat retina but not in the P23H rat retina. The photopic ERG of the dystrophic RCS rat retina becomes increasingly electronegative because of an aberrant negative response, originating from amacrine cell activity, which declines more slowly than the b-wave with degeneration. The absence of this response in the P23H rat indicates that the inner retinal cone pathway pathology is different in the two models. A relative increase in Kir4.1 channels on Müller cells of RCS retina may contribute to the enhanced negative ERG response in the RCS rat.

Morphological Evidence that Xenon Neuroprotects against N‐Methyl‐dl‐Aspartic Acid‐Induced Damage in the Rat Arcuate Nucleus

The hyperactivation of glutamate receptors, especially those of the N-methyl-d-aspartate subtype (NMDA), can induce excess calcium entry into cells, leading to neuronal death. Since the anesthetic gas xenon behaves as an NMDA antagonist, the present article investigated, by distinct morphological approaches and after different times, the possible neuroprotectant effects of this gas in a model of neuronal damage induced by N-methyl-dl-aspartic acid (NMA) on rat arcuate nucleus. Rats were assigned to the following groups: controls; xenon exposure; NMA treatment; or xenon exposure + NMA treatment. Animals were placed in an experimental cage and after 10 min a mixture of xenon (or nitrogen) 70% and oxygen 30% was delivered. After 3 h, 1, 2, 5, or 7 days from gas exposure, rats were euthanized and the whole brain was removed and processed for either transmission electron microscopy or light microscopy. In the arcuate nucleus from NMA-treated animals only 40-60% of cell population survived in all times with several degenerating neurons giving the typical appearance of a "bull's eye." At ultrastructural level, chromatin margination, nuclear shrinkage, mitochondria with matrix dilution, dilated endoplasmic cisternae, and electrondense cytoplasm were detected. Xenon alone did not induce changes, but reduced of about 50% NMA-induced cell loss as well as degenerating neurons, with the maximal neuroprotection at 7 days. These results confirm that in the rat arcuate nucleus NMA can induce a severe neuronal damage that is already marked after 3 h. Xenon significantly reduced the neuronal damage at all times and can be then regarded as a promising neuroprotectant agent.

Continuous Human Metastin 45–54 Infusion Desensitizes G Protein-Coupled Receptor 54-Induced Gonadotropin-Releasing Hormone Release Monitored Indirectly in the Juvenile Male Rhesus Monkey (Macaca mulatta): A Finding with Therapeutic Implications

The effect of continuous administration of the C-terminal fragment of metastin, the ligand for the G protein-coupled receptor, GPR54, on GnRH-induced LH secretion was examined in three agonadal, juvenile male monkeys whose responsiveness to GnRH was heightened by pretreatment with a chronic pulsatile iv infusion of synthetic GnRH. After bolus injection of 10 microg human (hu) metastin 45-54 (equivalent to kisspeptin 112-121), the GPR54 agonist was infused continuously at a dose of 100 microg/h and elicited a brisk LH response for approximately 3 h. This rise was then followed by a precipitous drop in LH despite continuous exposure of GPR54 to metastin 45-54. On d 4, during the final 3 h of the infusion, single boluses of hu metastin 45-54 (10 microg), N-methyl-DL-aspartic acid (NMDA) (10 mg/kg) and GnRH (0.3 microg) were administered to interrogate each element of the metastin-GPR54-GnRH-GnRH receptor cascade. Although the NMDA and GnRH boluses were able to elicit LH pulses, that of hu metastin 45-54 was not, demonstrating functional integrity of GnRH neurons (NMDA) and GnRH receptors (NMDA and GnRH) but desensitization of GPR54. The desensitization of GPR54 by continuous hu metastin 45-54 administration has therapeutic implications for a variety of conditions currently being treated by GnRH and its analogs, including restoration of fertility in patients with abnormal GnRH secretion (i.e. idiopathic hypogonadotropic hypogonadism and hypothalamic amenorrhea) and selective, reversible suppression of the pituitary-gonadal axis to achieve suppression of gonadal steroids (i.e. precocious puberty, endometriosis, uterine fibroids, and prostate cancer).

P.1.187 Chronic corticosterone attenuates 5-HT 1A-mediated but not GABA-mediated inhibition of rat dorsal raphe neurones

The effects of common antiepileptics, GABAmimetic drugs, excitatory amino acid antagonists as well as of clonidine, corynanthine, chlorpromazine and atropine were studied against clonic convulsion induced in mice by N-methyl-D,L-aspartic acid (NMDLA) after subcutaneous (340 mg/kg; ED97) administration. Among the antiepileptics studied, valproate (ED50: 340 mg/kg after subcutaneous injection of NMDLA) and diazepam (ED50: 0.78 mg/kg after intravenous and 14 mg/kg after subcutaneous injection of NMDLA) antagonized NMDLA-induced convulsions, whereas phenobarbital (up to 80 mgkg), diphenylhydantoin (up to 50 mg/kg) and ethosuximide (500 mg/kg) were totally ineffective. Moreover, 2-amino-5-phosphonopentanoic acid and 2-amino-7-phosphonoheptanoic acid (but not glutamic acid diethyl ester), aminooxyyacetic acid, cetyl GABA and clonidine protected strongly against the convulsant whereas progabide was only weakly active. THIP showed a higher activity against intravenous than against subcutaneous NMDLA. Baclofen and atropine even increased mortality and the remaining agents exerted no significant protective action. The data suggest that NMDLA-induced convulsions can be blocked effectively by direct antagonism of NMDLA-produced excitation, enhancement of GABA-mediated inhibition, and activation of central α2-adrenoceptors. The possible efficacy of valproate in cases of epilepsy with a distinct rise in plasma excitatory amino acid levels should be carefully considered.

Stimulation of luteinizing hormone secretion by N-methyl-D,L-aspartic acid in the adult male guinea-pig: incomplete blockade by gonadotropin-releasing hormone receptor antagonism.

Stimulation of luteinizing hormone (LH) secretion by N-methyl-D,L-aspartic acid (NMA), reported for several mammalian species, is generally accepted to be mediated through stimulation of hypothalamic gonadotropin-releasing hormone (GnRH) release. In view of a previously reported unexpected inhibitory action of NMA on GnRH release from hypothalamic explant of intact male guinea-pigs, the aim of the present study was to assess the in vivo effects of NMA in the adult male guinea-pig. In the gonadally intact male, NMA (5 mg/animal) elicited a robust LH secretion, which was blocked by the N-methyl-D-asparte-receptor antagonist DL-2-amino-5-phosphonovaleric acid (AP-5,12 mg/animal). In the castrated male, NMA elicited only a marginal and inconsistent LH secretion. Cetrorelix (CET), a GnRH receptor antagonist, administered intracardiacally 1 min or 45 min preceding bolus injection of NMA significantly reduced the LH response to NMA in the intact male. Surprisingly, following GnRH receptor blockade with CET, there still was a substantial residual serum LH response to NMA, while CET completely abolished the serum LH response to high dose (1 microg or 10 microg) guinea-pig GnRH (gpGnRH). These results indicate that NMA stimulates LH secretion in the gonadally intact male guinea-pig in vivo and that this effect is mediated in part through gpGnRH-independent mechanisms.

Neuroprotective and neurotoxic properties of the ‘inert’ gas, xenon

Antagonists of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors have been shown not only to have neuroprotective effects but also to exhibit neurotoxic properties. In this study, we used c-Fos, a protein product of an immediate early gene, as a marker of neuronal injury to compare the neuroprotective effects of xenon and the neurotoxic properties of xenon, nitrous oxide, and ketamine, three anaesthetics with NMDA receptor antagonist properties.We used an in vivo rat model of brain injury in which N-methyl-DL-aspartic acid (NMA) is injected subcutaneously (s.c.) and c-Fos expression in the arcuate nucleus is used as a measure of injury. To examine the neurotoxic potential of each of the three anaesthetics with NMDA receptor antagonist properties, c-Fos expression in the posterior cingulate and retrosplenial (PC/RS) cortices was measured.Xenon dose-dependently suppressed NMA-induced c-Fos expression in the arcuate nucleus with an IC(50) of 47 (2)% atm. At the highest concentration tested (75% atm) NMA-induced neuronal injury was decreased by as much as that observed with the prototypical NMDA antagonist MK801 (0.5 mg kg(-1) s.c.). Both nitrous oxide and ketamine dose-dependently increased c-Fos expression in PC/RS cortices; in contrast, xenon produced no significant effect. If the dopamine receptor antagonist haloperidol was given before either nitrous oxide or ketamine, their neurotoxic effects were eliminated.Uniquely amongst anaesthetics with known NMDA receptor antagonist action, xenon exhibits neuroprotective properties without co-existing neurotoxicity. The reason why ketamine and nitrous oxide, but not xenon, produce neurotoxicity may involve their actions on dopaminergic pathways.

Neuroprotective and neurotoxic properties of the 'inert' gas, xenon

Antagonists of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors have been shown not only to have neuroprotective effects but also to exhibit neurotoxic properties. In this study, we used c-Fos, a protein product of an immediate early gene, as a marker of neuronal injury to compare the neuroprotective effects of xenon and the neurotoxic properties of xenon, nitrous oxide, and ketamine, three anaesthetics with NMDA receptor antagonist properties. We used an in vivo rat model of brain injury in which N-methyl-DL-aspartic acid (NMA) is injected subcutaneously (s.c.) and c-Fos expression in the arcuate nucleus is used as a measure of injury. To examine the neurotoxic potential of each of the three anaesthetics with NMDA receptor antagonist properties, c-Fos expression in the posterior cingulate and retrosplenial (PC/RS) cortices was measured. Xenon dose-dependently suppressed NMA-induced c-Fos expression in the arcuate nucleus with an IC(50) of 47 (2)% atm. At the highest concentration tested (75% atm) NMA-induced neuronal injury was decreased by as much as that observed with the prototypical NMDA antagonist MK801 (0.5 mg kg(-1) s.c.). Both nitrous oxide and ketamine dose-dependently increased c-Fos expression in PC/RS cortices; in contrast, xenon produced no significant effect. If the dopamine receptor antagonist haloperidol was given before either nitrous oxide or ketamine, their neurotoxic effects were eliminated. Uniquely amongst anaesthetics with known NMDA receptor antagonist action, xenon exhibits neuroprotective properties without co-existing neurotoxicity. The reason why ketamine and nitrous oxide, but not xenon, produce neurotoxicity may involve their actions on dopaminergic pathways.

Nitric oxide regulation of gonadotrophin secretion in prepubertal heifers1.

Mechanisms responsible for the pulsatile release of gonadotrophin secretion in prepubertal heifers are not fully known. We have shown that an excitatory amino acid agonist, N-Methyl-D,L-aspartic acid (NMA), induces an immediate release of luteinizing hormone (LH) and follicle stimulating hormone (FSH) in prepubertal heifers. Nitric oxide (NO) has also emerged as an important regulator of LH release in rats. This study was designed to test the role of NO in the regulation of gonadotrophin release as well as the possible mediation by NO of the effects of NMA and gonadotrophin releasing hormone (GnRH) on gonadotrophin secretion in heifer calves. In experiment 1, four groups of five prepubertal heifers (33 weeks old) received one of the following treatments: (1); N-G-nitro-L-arginine methyl ester (L-NAME, a NO synthase inhibitor, 35 mg/kg, i.v., once); (2) NMA (4.7 mg/kg, i.v., once); (3) L-NAME+NMA (as above); and (4) Vehicle (saline, i.v.). All heifers in all groups were also challenged with a bolus injection of GnRH (10 ng/kg, i.v., once). Blood samples were collected every 15 min for 10 h. L-NAME was injected after the first blood sample, NMA after 2 h and GnRH after 6 h of blood sampling. Administration of L-NAME alone, suppressed the spontaneous pulses of LH (P<0.04). Heifers in the NMA group responded with a significantly greater LH release than did the heifers in the L-NAME+NMA group (P<0.05). Following the GnRH challenge, heifer calves treated with L-NAME or NMA had higher LH pulse responses than the controls (P<0.05). In a second experiment, four groups of five heifer calves (34 weeks old) were given one of the following treatments: (1) L-NAME (as above); (2) L-arginine, a NO precursor (ARG, 100 mg/kg/h, i.v. drip infused for 6 h starting 2 h after first blood sample was taken); (3) L-NAME+ARG (as above); and (4) Vehicle (saline i.v. bolus and drip for 6 h). Blood samples were taken every 10 min for 8 h. Administration of L-NAME suppressed the pulsatile release of LH and FSH (P<0.05). Compared to the control group, infusion of ARG by itself did not change the pattern of LH secretion (P>0.05); however, in heifers given L-NAME, ARG restored a normal pattern of LH pulses, similar to the control values (P>0.05). It was therefore concluded that NO is involved in the regulation of LH, and possibly FSH, secretion and that NO may mediate, at least in part, the stimulatory effects of NMA on LH, and to some extent FSH, release. The responses to GnRH led us to suggest that NO may have inhibitory effects on the pituitary and NMA may have increased pituitary sensitivity to GnRH.

Testosterone modulates growth hormone secretion at the hypothalamic but not at the hypophyseal level in the adult male rhesus monkey

We investigated a possible modulation of growth hormone (GH) secretion by testosterone by measuring the growth hormone releasing hormone (GHRH)-stimulated and N-methyl-d,l-aspartic acid (NMA)-induced GH secretion in adult rhesus monkeys. Intact, orchidectomized and testosterone-substituted (testosterone enanthate 125 mg/week, i.m. for 5 weeks) orchidectomized monkeys (n=5) were used in the study. GHRH (25 microg/kg body weight) or NMA (15 mg/kg body weight) was infused through a Teflon cannula implanted in the saphenous vein. Sequential blood samples were collected 30-60 min before and 60 min after the injection of the neurohormone or the drug at 10-20-min intervals. All bleedings were carried out under ketamine hydrochloride anaesthesia (initial dose 5 mg/kg body weight i.m., followed by 2.5 mg/kg at 30-min intervals). The plasma concentrations of GH, testosterone and oestradiol (E(2)) were determined by using specific assay systems. Administration of GHRH elicited a significant increase in GH secretion in all three groups of animals. There was no significant difference in the responsiveness of pituitary somatotrophs to exogenous GHRH challenges between intact and orchidectomized monkeys and testosterone replacement in orchidectomized animals did not significantly alter the GHRH-induced GH response. The responsiveness of hypothalamic GHRH neurones apparently did undergo a qualitative change after orchidectomy, as GH response to NMA was less in orchidectomized animals than in intact monkeys. The responsiveness of GHRH neurones to exogenous NMA was restored and even potentiated when orchidectomized monkeys were treated with testosterone. Taken together, these findings suggest that testosterone does not affect the sensitivity of the pituitary somatotrophs to GHRH but stimulates the secretion of GH by modulation of the NMDA drive to GHRH neurones.

Excitatory amino acid regulation of gonadotropin secretion in prepubertal heifer calves.

The mechanisms controlling the pulsatile release of gonadotropins in prepubertal heifers are not completely understood. We examined the role of excitatory amino acid neurotransmitters, via activation of N-methyl-D-aspartate (NMDA) receptors, in the control of pulsatile LH and FSH release during prepubertal development in heifers. Hereford heifer calves received 4.7 mg/kg of N-methyl-D,L-aspartic acid (NMA), a potent NMDA receptor agonist (n = 5, i.v.), or saline (n = 5, i.v.), as single doses, at 4, 8, 12, 24, 36, and 48 wk of age. Blood samples were collected every 15 min, for 1 h before and 9 h after injection, on the days of treatment. Injection of NMA resulted in an acute release of LH (p < 0.001) in 0, 3, 3, 4, 5, and 5 calves (p < 0.01) and of FSH (p < 0.001) in 0, 1, 2, 4, 3, and 2 calves at 4, 8, 12, 24, 36, and 48 wk of age, respectively. The peak response of LH and FSH release to NMA was at 15 min posttreatment, and these peak responses were highest at 36 wk of age (p < 0.05). We suggest that neuroexcitatory amino acids, through NMDA receptors, are involved in prepubertal development of LH and FSH secretion in heifer calves.

Premature elevation in serum insulin-like growth factor-I advances first ovulation in rhesus monkeys.

Previous data suggest that developmental increases in peripheral concentrations of insulin-like growth factor-I (IGF-I) may be one of several neuroendocrine signals that regulate the timing of puberty. In order to test this hypothesis further, normal juvenile female rhesus monkeys (Con; n = 6) were compared with age-matched animals (Igf; n = 4) which received a constant subcutaneous infusion of recombinant human IGF-I (110 micrograms/kg/day) from 18 through 36 months of age. Menstrual bleeding was monitored and ovulation was inferred from a sustained rise in serum progesterone. In order to assess the sensitivity of luteinizing hormone-releasing hormone (LHRH) neurons to excitation, the response of serum LH to the acute administration of the glutamate receptor agonist N-methyl-D, L-aspartic acid (NMDA) was assessed prior to menarche, 2 months following menarche, and during the follicular phase of a female's third ovulation or 50 days after a female's first ovulation. In addition, the pituitary response of LH secretion to an LHRH agonist was assessed during the follicular phase of a female's fourth ovulation or 75 days following her first ovulation. IGF-I treatment effectively elevated serum concentrations by more than 86% of the values observed in Con animals. Although the treatment also enhanced the developmental increase in IGF binding protein-3 (IGFBP-3), IGF-I was increased proportionately more, resulting in a significantly higher molar ratio of IGF-I:IGFBP-3 in treated females throughout the course of the study. Treatment with IGF-I did not affect age at menarche but did significantly advance the age of first ovulation. Consequently, the interval between menarche and first ovulation was significantly shorter in Igf compared with Con females. Although the total number of ovulations exhibited by Igf (3.8 +/- 0.3) and Con females (3.0 +/- 0.5) in the 12 months following menarche was similar, significantly more of these were characterized by normal luteal phase progesterone secretion in Igf (100% +/- 0) compared with Con females (64% +/- 17). An analysis of cycles with a normal luteal phase revealed that serum estradiol during the luteal phase was significantly higher in Igf compared with Con females. Finally, IGF-treated females responded to NMDA treatment with a significantly greater increase in serum LH following menarche but not prior to menarche. In contrast, the response of serum LH to an LHRH agonist did not differ between Igf and Con females. These data suggest that the premature elevation in IGF-I levels, and consequently the ratio of IGF-I:IGFBP-3, accelerates the tempo of the final stages of puberty in rhesus monkeys. This action of IGF-I is probably the result of an increase in LHRH neuronal activity and is not due to a change in pituitary sensitivity to LHRH stimulation. In addition, ovarian sensitivity to LH stimulation during the luteal phase is also increased by IGF-I. Taken together, these data suggest that developmental increases in peripheral IGF-I secretion participate in the neuroendocrine regulation of puberty in female primates.

Antimicrobial and anticonvulsant activities of Viscum capense

Dichloromethane, methanol and water extracts of Viscum sapense L.f., of the Loranthaceae family, were tested for antimicrobial activities against Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans. Methanol extract was also tested for activity against seizures in albino mice induced by pentylenetetrazole (PTZ), bicuculline and N-methyl- dl-aspartic acid (NMDLA). Methanol extract of V. capense inhibited the growth of S. aureus. Methanol extract also protected the mice against PTZ- and bicuculline-induced tonic seizures but did not significantly alter NMDLA-induced tonic seizures. The data indicate that the extract of V. capense has antibacterial activity against S. aureus and also anticonvulsant activity.