Biomedical subjects

S W Leslie

Publications and source records attributed to S W Leslie.

At least 19 recordsLinked to original sources

PubMed2003

Acute ethanol affects phosphorylation state of the NMDA receptor complex: implication of tyrosine phosphatases and protein kinase A.

Phosphorylation has been shown to regulate N-methyl-D-aspartic acid receptor (NMDAR) function. The inhibitory effect of ethanol on NMDAR function could be due, at least in part, to a change in NMDAR phosphorylation states. In order to investigate the effect of ethanol on phosphorylation of NR1 and NR2 subunits, NMDAR complexes were immunoprecipitated from cortical slices pre-exposed to ethanol. Acute ethanol, 100 and 200 mM, significantly decreased the tyrosine phosphorylation of NR2 subunits (Tyr-NR2). Treatment with a tyrosine phosphatase inhibitor reduced the inhibition of Tyr-NR2 phosphorylation caused by 100 mM ethanol. This suggests an involvement of tyrosine phosphatases in ethanol-induced inhibition of Tyr-NR2 phosphorylation. Slices pre-exposed to 100 and 200 mM ethanol exhibited a significant increase in the phosphorylation of NR1 by PKA at serine 897 (Ser897-NR1), which was blocked by a PKA inhibitor. Moreover, at 200 mM, ethanol produced a significant increase in PKA activity. Together, these results indicate that ethanol may increase Ser897-NR1 phosphorylation by activating PKA. However, ethanol did not affect phosphorylation of NR1 subunits by PKC at serine 896. We conclude that ethanol has the ability to modulate phosphorylation of both NR2 and NR1 subunits and these effects appear to implicate tyrosine phosphatases and PKA, respectively.

Alcohol-Induced Disorders, Nervous System↗

PubMed2003

Cell surface expression of NR1 splice variants and NR2 subunits is modified by prenatal ethanol exposure.

N-methyl-D-aspartate receptor dysfunction has been strongly suggested to link with the abnormalities seen in fetal alcohol syndrome. Thus, the effects of prenatal ethanol exposure on the total expression of NR1 splice variants and the cell surface expression of both NR1 and NR2 subunits in brain were investigated in rats. Western blot studies of membrane homogenates from cerebral cortices at postnatal days 1 through 21 indicate that prenatal ethanol treatment does not alter total NR1 expression or differential expression of NR1 splice variants during development. However, immunoprecipitation studies using PSD95 suggest that both C2'-terminal variants and NR2A subunits at the cortical postsynaptic membrane of postnatal day 21 were significantly reduced after prenatal ethanol treatment. Moreover, C1-terminal variants were decreased in both pair-fed and ethanol-treated groups, while no significant differences in the levels of total NR1 subunits, NR1 splice variants containing the N- or C2-terminal cassettes, or NR2B subunits were observed. Thus, these results suggest that prenatal exposure to ethanol may influence neuronal function by selective regulation of expression of C2'-terminal variants and NR2A subunits at the cell surface.

Aging↗

PubMed2002

DARPP-32 and regulation of the ethanol sensitivity of NMDA receptors in the nucleus accumbens.

The medium spiny neurons of the nucleus accumbens receive both an excitatory glutamatergic input from forebrain and a dopaminergic input from the ventral tegmental area. This integration point may constitute a locus whereby the N-methyl-D-aspartate (NMDA)-subtype of glutamate receptors promotes drug reinforcement. Here we investigate how dopaminergic inputs alter the ethanol sensitivity of NMDA receptors in rats and mice and report that previous dopamine receptor-1 (D1) activation, culminating in dopamine and cAMP-regulated phosphoprotein-32 kD (DARPP-32) and NMDA receptor subunit-1 (NR1)-NMDA receptor phosphorylation, strongly decreases ethanol inhibition of NMDA responses. The regulation of ethanol sensitivity of NMDA receptors by D1 receptors was absent in DARPP-32 knockout mice. We propose that DARPP-32 mediated blunting of the response to ethanol subsequent to activation of ventral tegmental area dopaminergic neurons initiates molecular alterations that influence synaptic plasticity in this circuit, thereby promoting the development of ethanol reinforcement.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗

PubMed2001

Effect of gestational ethanol exposure on the NMDA receptor complex in rat forebrain: from gene transcription to cell surface.

Effects of gestational ethanol exposure on the trafficking of the NMDA receptor complex were investigated. Studies focused on three distinct processes in NMDA receptor translocation: (1) the level of gene transcription (2) nascent NMDA receptor subunits (NR) associated with the endoplasmic reticulum bound chaperone protein calnexin and (3) NMDA receptors associated with the cell surface anchoring protein PSD-95. Forebrain mRNA and membrane proteins were isolated from postnatal day 1 rat pups from prenatally ethanol exposed, pair-fed and ad libitum experimental groups. Ribonuclease protection assays were carried out to determine the levels of NR2A, NR2B, and NR2C mRNA within the treatment groups determined. Results indicated that gestational ethanol exposure did not affect the gene transcription of the NR2 subunits. Immunoprecipitation experiments were conducted with an anti-calnexin antibody or an anti-PSD-95 antibody and the immunoprecipitates probed for NR1 and NR2 subunits. Within the anti-calnexin immunoprecipitates, no NR2A, NR2B or NR2C subunits were detectable, but a significant pool of NR1 subunits was identified. These findings suggest that NR1 subunits but not NR2 subunits are associated with calnexin within the endoplasmic reticulum. Further, gestational ethanol exposure significantly increased the NR1 polypeptide levels in the anti-calnexin immunoprecipitate. Anti-PSD-95 immunoprecipitates revealed an abundance of NR1 and NR2B subunits, and these complexes were unaffected by gestational ethanol exposure. No NR2A or NR2C subunits were detected. These results suggest that gestational ethanol exposure significantly affects the assembly and transport of NMDA receptors. Gestational ethanol exposure may not alter the composition of the PSD-95 associated NMDA receptor complex.

Animals↗

PubMed2001

Acute exposure of cerebellar granule neurons to ethanol suppresses stress-activated protein kinase-1 and concomitantly induces AP-1.

The current studies were designed to examine the mechanisms of acute effects of ethanol on cerebellar granule neurons (CGNs) during neurodevelopment, with specific reference to activator protein-1 (AP-1). CGNs, isolated from 3-day-old Sprague-Dawley rats and cultured for 3 days, were exposed to 0, 22.5, and 100 mM ethanol for 1 h. Gel shift assays performed on the nuclear protein extracts showed increased AP-1 and heat shock factor-1 (HSF-1) transcriptional activation in response to ethanol. Western blots and RT-PCR showed increased c-JUN and phosphorylated c-JUN (serine 73) protein, as well as c-jun mRNA. Ethanol paradoxically decreased the activity of stress-activated protein kinase-1 (SAPK-1) while increasing p44 and p42 mitogen-activated protein kinase (MAPK) activity. The protein synthesis-inhibiting and SAPK-1 activity-inducing antibiotic, anisomycin (30 and 500 microM) decreased AP-1 transcriptional activation to 47 and 23% of control values, respectively. The anisomycin effect was enhanced in the presence of 100 mM ethanol. Similarly, cycloheximide decreased ethanol-induced AP-1 transcriptional activation. Pretreatment with the MAPK kinase (MEK) pathway inhibitor PD98059 resulted in decreases in both ethanol-induced and control AP-1 DNA binding. Thus this acute ethanol-induced increased AP-1 transcriptional activation requires protein synthesis and involves MEK-independent increased MAPK phosphorylation, on the one hand, and decreased SAPK-1 activity on the other. The ethanol effect is thus ascribed to the activities of alternate kinase pathways and/or the inhibition of (a) protein phosphatase(s). Exposure of CGNs to ethanol for 24 h resulted in decreased AP-1 DNA binding, an observation that could have consequences for overall neuronal function under chronic exposure conditions.

Animals↗

PubMed2000

Impact of body weight on urinary electrolytes in urinary stone formers.

OBJECTIVES: Obesity increases the risk of developing chronic medical conditions such as diabetes mellitus, hypertension, and coronary artery disease. We performed a retrospective review of a large data base on urinary stones to determine if differences are found in urine and serum chemistries among obese and nonobese stone-forming patients. The effect of body weight on stone recurrence among urinary stone formers was also determined. METHODS: A national data base containing serum biochemical profiles, 24-hour urine specimens, and standardized questionnaires was retrospectively evaluated from 5942 consecutive patients with urinary stone disease. Stone-forming patients were classified by body weight: nonobese men, less than 100 kg and nonobese women, less than 85 kg; intermediate men, 100 to 120 kg and intermediate women, 85 to 100 kg; and obese men, more than 120 kg and obese women, more than 100 kg. RESULTS: Obese stone formers comprised 6.8% (n = 404) of the patient population. The mean weight in the nonobese and obese groups was 81 kg versus 134 kg, respectively, for men and 64 kg versus 112 kg, respectively, for women. Obese patients represented 3.8% of the male and 12.6% of the female population. Obese patients had increased urinary excretion of sodium, calcium, magnesium, citrate, sulfate, phosphate, oxalate, uric acid, and cystine; obesity was associated with increased urinary volumes and urine osmolality compared with the nonobese patients. Obese men had increased concentration of urinary sodium, oxalate, uric acid, sulfate, and phosphate when corrected for urinary volume. Obese women had increased concentrations of sodium, uric acid, sulfate, phosphate, and cystine. The mean number of stone episodes in nonobese versus obese men was similar (3.55 and 3.56), whereas mean stone episodes were 2.93 and 3.38 (P = 0.045) for nonobese versus obese women. CONCLUSIONS: Among known stone formers, obesity is associated with unique changes in both serum and urinary chemistries. These changes are associated with an increased incidence of urinary stone episodes in obese women but not in obese men.

Adolescent↗

PubMed1998

Effect of prenatal ethanol exposure on the developmental profile of the NMDA receptor subunits in rat forebrain and hippocampus.

The effects of prenatal ethanol exposure on the NMDAR1 protein expression (postnatal days 1 and 7) and on the developmental profile of the NMDAR2A and NMDAR2B subunits in rat forebrain and hippocampus were investigated. Forebrain and hippocampal membrane proteins were isolated from pups of various ages (postnatal days 1 to 21) from prenatally ethanol exposed, pair-fed and ad libitum control groups. A semiquantitative immunoblot procedure was used with antibodies raised against the NMDAR1, NMDAR2A, and the NMDAR2B subunits to assess the NMDA subunit protein expression in the samples. NMDAR1 protein expression was unaffected by prenatal ethanol exposure at postnatal day 1 or 7 in both the forebrain and hippocampus. NMDAR2A protein expression levels rose rapidly in both forebrain and hippocampus during the time frame of study. Prenatal ethanol exposure caused a significant reduction in protein expression levels of the NMDAR2A in forebrain through postnatal day 14. NMDAR2B protein expression levels were high throughout the study in both forebrain and hippocampus. Prenatal ethanol exposure significantly reduced protein expression of the NMDAR2B in the forebrain (through postnatal day 14) and hippocampus (up to day 7). The results suggest that there may be a link between the depressed expression of the NMDAR2 subunits and the neurodevelopmental disorders associated with fetal ethanol exposure.

Animals↗

PubMed1997

A partial agonist model used in the allosteric modulation of the NMDA receptor.

We used a partial agonist model to understand further the allosteric modulation of D,L-(E)-2-amino4-propyl-5-phosphono-3-pentenoic acid ([3H]CGP-39653) binding by glycine, 1-hydroxy-3-amino-2-pyrrolidone (HA-966) and 5,7-dichlorokynurenic acid at the NMDA receptor. Binding of [3H]CGP-39653 was investigated in homogenates of cortex, hippocampus and cerebellum of adult rat. Glycine, HA-966 and 5,7-dichlorokynurenic acid maximally decreased the binding of 10 nM of [3H]CGP-39653 by approximately 50, 40 and 22%, respectively. Glycine, HA-966 and 5,7-dichlorokynurenic acid reduced [3H]CGP-39653 binding with IC50 values of 0.31, 11 and 0.044 microM, respectively. The decrease in [3H]CGP-39653 binding was due to a reduced affinity (Kd) and number of binding sites (Bmax) by all three drugs at concentrations where approximately maximum inhibition was observed. Glycine, HA-966 and 5,7-dichlorokynurenic acid lowered the Bmax by approximately 29, 16 and 10%, respectively, whereas the Kd values were increased by approximately 84, 44 and 32%, respectively, in cortex and hippocampus. There was no change in the binding of [3H]CGP-39653 in the cerebellum. The model used revealed that neither 5,7-dichlorokynurenic acid nor HA-966 had partial agonist characteristics in respect with the allosteric modulation of [3H]CGP-39653 binding. Furthermore, the results showed that brain regions have different pharmacological profiles which may depend on the NMDA receptor subunit composition.

2-Amino-5-phosphonovalerate↗

PubMed1997

Geriatric urolithiasis.

PURPOSE: We define the differences between geriatric patients with urinary stone disease compared to a younger cohort. MATERIALS AND METHODS: A data base, including serum biochemical profiles, 24-hour urinalyses and standardized questionnaires, was retrospectively evaluated from more than 6,000 consecutive patients with urinary stone disease. RESULTS: Geriatric stone formers comprised 12% (721) of all stone patients. Two-thirds of these elderly patients had aberrant urinary values and 29% had isolated hypocitraturia compared to 17% in the younger group. Of geriatric stone forming patients 76% had recurrent urinary stones (mean 3.5 stone episodes), which was similar to the younger comparable group (77%, mean 3.3 stone episodes). The severity of urinary stone disease was similar between the 2 groups based on the need for urological intervention. Geriatric stone patients, in general, experienced the first stone episode later in life (after age 50 years) compared with younger patients. Elderly patients had an increased incidence of uric acid stones, but had a similar incidence of struvite calculi. Geriatric stone patients underwent parathyroid surgery more frequently (2.7 versus 0.7%). Geriatric stone forming patients rarely had renal failure. CONCLUSIONS: The incidence, recurrence and severity of recurrent urinary stone disease were similar between geriatric and younger stone forming patients. Geriatric stone patients had an increased incidence of isolated hypocitraturia, uric acid calculi and previous parathyroidectomy. The geriatric stone population is not merely an extension of younger stone forming patients presenting at an older age. Rather, geriatric patients commonly experience the first symptomatic stone episode later in life.

Age Distribution↗

PubMed1997

Effects of prenatal ethanol exposure on brain region NMDA-mediated increase in intracellular calcium and the NMDAR1 subunit in forebrain.

The effects of prenatal ethanol exposure on N-methyl-D-aspartate (NMDA)-mediated calcium entry into neonatal dissociated neurons from hippocampus, forebrain, and cerebellum were investigated. Dissociated cells were isolated from less than 1-day-old pups of prenatally exposed, pair-fed control and ad libitum control groups and loaded with fura-2. Prenatal ethanol exposure significantly reduced the NMDA-stimulated increase in intracellular calcium in all three brain regions compared to the two control groups. These findings are very similar to those previously observed in neonatal dissociated whole brain neurons using the same ethanol exposure protocol. Studies were also conducted using forebrain to determine if prenatal ethanol exposure alters NMDAR1 subunit protein expression in this major brain area; however, the results indicated no significant differences between ethanol-exposed and control groups.

Animals↗

PubMed1997

Effects of prenatal and early postnatal ethanol exposure on [3H]MK-801 binding in rat cortex and hippocampus.

The effects of prenatal and/or early postnatal exposure to ethanol at high concentrations on N-methyl-D-aspartate (NMDA) receptor number and functioning in the weanling rat were examined. The binge-like exposure protocol was used in an animal model of acute ethanol effects at two critical periods of development. [3H]MK-801 binding parameters for the internal channel phencyclidine site were assessed in the presence of 10 microM glutamate and 10 microM glycine activation. Four treatment groups were included: (1) animals exposed to ethanol both prenatal and postnatal; (2) animals exposed only prenatal; (3) animals exposed early postnatal only; and (4) control animals with no exposure to ethanol. The results of the [3H]MK-801 binding experiments showed that both prenatal and postnatal exposure to ethanol resulted in a significant decrease in the density of NMDA receptors. In addition, data indicated an apparent increase in the percentage of high-affinity state (open channel state) relative to low-affinity state (close channel state) receptors in the ethanol-treated groups. These results show that both prenatal and postnatal ethanol exposure decrease NMDA receptor density in the cortex and hippocampus. The findings are consistent with previous observations by our laboratory and others that NMDA-mediated calcium influx is reduced in these regions, as well as in whole brain by prenatal ethanol exposure. It is suggested that after ethanol exposure, the remaining functional NMDA receptors might have altered sensitivity to coagonist activation with an increased probability of channel opening.

Animals↗

PubMed1996

N-methyl-D-aspartate-stimulated increases in intracellular calcium exhibit brain regional differences in sensitivity to inhibition by ethanol.

This study compared N-methyl-D-aspartate (NMDA)-stimulated increases in intracellular calcium in fura-2-loaded neurons dissociated from newborn rat brainstem (EC50 in microM; 6.4), cerebellum (9.5), forebrain (6.3), and hippocampus (10.6). Ethanol inhibition of the response to 25 microM NMDA differed among the regions. The NMDA response in hippocampus was inhibited by 20 mM ethanol; cortex and cerebellum responses were inhibited by 80 mM ethanol, and no inhibition was seen in the brainstem. Addition of glycine (15 microM) failed to attenuate ethanol inhibition of the NMDA response. These results demonstrate that ethanol inhibition of NMDA-stimulated responses varies according to brain region. In contrast to previous findings from this laboratory using dissociated neurons from whole brain, the addition of glycine did not reverse the inhibitory effects of ethanol on NMDA-stimulated responses.

Animals↗

PubMed1995

Alteration of [3H]MK-801 binding associated with the N-methyl-D-Aspartate receptor complex by acute ethanol in rat cortex and hippocampus in vitro.

We investigated the effect of ethanol on specific binding of [3H]MK-801 to the intrachannel phencyclidine (PCP) receptor site, as an index of change in the functional response of the N-methyl-D-Aspartate (NMDA)-associated ion channel. Saturation binding experiments were performed on synaptic membrane homogenates from adult rat cortex and hippocampus. [3H]MK-801 binding assays were conducted under conditions of basal, 10 microM glutamate, or 10 microM glutamate + 30 microM D-serine, with and without 50 or 100 mM ethanol. Association experiments of [3H]MK-801 binding (5 nM) were conducted under conditions of 0 or 10 microM glutamate, with varying concentrations of glycine (0.01, 0.10, and 10 microM) with and without 100 mM ethanol. Ethanol (50 and 100 mM) significantly decreased the percentage of high-affinity (open-channel state) MK-801 receptors with a concomitant increase in percentage of low-affinity receptors, but did not change high- and low-affinity constants of the two binding states. An ethanol-induced increase in the closed-channel receptor density in basal and activated conditions was suggested by the saturation experiments. Association experiments further explained this finding, in that ethanol (100 mM) significantly decreased fast component (open-channel) [3H]MK-801 binding in conditions of glycine (0.01-10 microM) only and activated conditions of glutamate + glycine (0.01-0.10 microM). However, the observed fast and slow kinetic rate constants of [3h]MK-801 binding, as well as total specific binding (fast + slow components), were not altered.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

Effects of prenatal ethanol exposure on N-methyl-D-aspartate-mediated calcium entry into dissociated neurons.

The effects of prenatal ethanol exposure on N-methyl-D-aspartate (NMDA)-activated calcium entry into dissociated neurons were studied. Dissociated brain cells were isolated from less than 1-day-old pups from prenatally ethanol-exposed, pair-fed control and ad libitum control groups and loaded with fura-2. Prenatal ethanol exposure significantly decreased the NMDA receptor-mediated calcium entry compared to both pair-fed and ad libitum control groups. To determine the mechanisms of the prenatal ethanol exposure on the NMDA-mediated ion channel decrements, possible modulatory sites of the NMDA receptor were studied. Glycine (0.1, 1, 10 and 100 microM) increased calcium entry to an equal extent in the ethanol and control groups, but did not reverse the effect of prenatal ethanol exposure. Furthermore, low concentrations of MK801 [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]-cyclohepten-5-10-imine hydrogen maleate] (25 and 50 nM) did not further inhibit calcium entry beyond that observed with the prenatal ethanol exposure, but significantly inhibited control group responses. Mg++ showed a similar result. With increasing concentrations of Mg++ the calcium entry in the three groups tended to converge. Thus, these results suggest that prenatal ethanol exposure inhibits the function of NMDA receptor-mediated ion channels by possibly altering the structural properties of the ion channel itself and/or by interacting with inner ion channel modulatory sites activated by Mg++ or MK801, and not the glycine site.

Animals↗

PubMed1993

Effects of in vitro ethanol and fetal ethanol exposure on glutathione stimulation of N-methyl-D-aspartate receptor function.

The present studies investigated the effects of glutathione (GSH; gamma-glutamylcysteinylglycine) and its oxidized form (GSSG) on neuronal N-methyl-D-aspartate (NMDA) receptor activation in both acute and chronic preparations of ethanol exposure. It was demonstrated using fura-2-loaded dissociated brain cells from newborn rat pups that both GSH and GSSG (0-4 mM) produced concentration-dependent increases in intracellular calcium similar to those produced by NMDA and other agonists of the NMDA receptor. GSH-stimulated calcium entry was not inhibited by low intoxicating concentrations of ethanol, which contrasts with ethanol's typical inhibitory effect on NMDA-stimulated receptor activation. Behavioral studies in adult rats demonstrated that ethanol-induced sleep times were significantly decreased when 10 microliters of GSSG (20 mM) were administered intracerebroventricularly approximately 5 min before an intraperitoneal injection of 20% (w/v) ethanol (3 g/kg). These findings suggest that the less potent effect of ethanol on GSH-stimulated calcium entry as well as the reduction in ethanol-induced sleep times may be related to the presence of glycine in the peptide. The glycine found in GSH may activate the glycine site and block or reduce ethanol's action on this site. It appears that although GSH may play an important role in the activation of the NMDA receptor, this action does not involve a process that is sensitive to acute ethanol exposure. In contrast, when rat pups were chronically exposed to ethanol via prenatal exposure before the fura-2 preparation, increases in NMDA- and GSH-stimulated calcium entry were significantly decreased relative to those in pair-fed and ad libitum-fed controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Stimulation of N-methyl-D-aspartate receptor-mediated calcium entry into dissociated neurons by reduced and oxidized glutathione.

The effects of GSH (gamma-glutamylcysteinylglycine) and GSSG on intracellular calcium levels ([Ca2+]i) were investigated using fura-2-loaded dissociated brain cells from newborn rat pups. Both produced concentration-dependent increases in [Ca2+]i (EC50 values of 914.3 +/- 190.5 and 583.0 +/- 97.2 microM for GSH and GSSG, respectively), similar to that observed with N-methyl-D-aspartate (NMDA) and other agonists at the NMDA receptor. Maximum response (expressed as percentage change in [Ca2+]i relative to basal) was significantly greater for GSSG (37.5 +/- 1.6%) than for GSH (25.3 +/- 1.6%). The response to both agents was prevented or reversed by competitive (100 microM) (-)-2-amino-5- phosphonovalerate and noncompetitive (400 nM) MK-801 or 1.0 mM Mg2+ antagonists of NMDA receptor-mediated calcium entry, even at concentrations of GSH and GSSG normally producing maximal response. The idea that these effects are mediated, at least in part, by interaction with the NMDA receptor was supported by the effects of GSH and GSSG on the binding of the NMDA receptor ligand [3H]CGP-39653 to membranes isolated from hippocampal and cortical homogenates. Both GSH and GSSG displaced bound [3H]CGP-39653, with IC50 values of 0.93 +/- 0.18 and 11.02 +/- 1.22 microM, respectively, and produced an increase in the apparent Kd of binding (control, 8.92 +/- 0.83 nM, and GSH, 13.31 +/- 1.19 nM; control, 11.59 +/- 0.35 nM, and GSSG, 18.73 +/- 0.66 nM). However, both also produced modest reductions in Bmax (control, 1265 +/- 69 fmol/mg of protein, and GSH, 901 +/- 73 fmol/mg of protein; control, 1068 +/- 30 fmol/mg of protein, and GSSG, 730 +/- 18 fmol/mg of protein) and Hill slopes (GSH, 0.66 +/- 0.02; GSSG, 0.62 +/- 0.04). This suggests complex kinetics for the interaction of GSH and GSSG with the NMDA receptor. Taken together, the results suggest the potential for modulation of the NMDA receptor complex by GSH and GSSG.

2-Amino-5-phosphonovalerate↗

PubMed1991

The effects of chronic ethanol exposure on N-methyl-D-aspartate-stimulated overflow of [3H]catecholamines from rat brain.

The effects of acute and chronic ethanol administration on N-methyl-D-aspartate-stimulated catecholamine overflow were examined. Three groups of male Sprague-Dawley rats were used. The first group received a chronic liquid diet containing ethanol (37%) for 3 weeks. The second group was pair-fed a liquid diet with dextrin substituted for ethanol isocalorically. The third group received Purina rat lab chow and water ad libitum. N-methyl-D-aspartate-stimulated [3H]catecholamine overflow from brain tissue slices was determined. N-methyl-D-aspartate (50-2000 microM) produced a concentration-dependent increase in [3H]norepinephrine overflow from cortical and hippocampal slices with no significant alteration of the response following chronic ethanol treatment. [3H]Dopamine overflow from striatal slices of the chronic ethanol group was significantly different at 1000 microM N-methyl-D-aspartate. The response of the chronic ethanol-treated group at the 1000 microM N-methyl-D-aspartate concentration was 30% and 40% lower than the pair-fed and ad libitum controls, respectively. Ethanol when added in vitro (30-200 mM) produced a concentration-dependent inhibition of N-methyl-D-aspartate (150 microM) stimulated efflux in all brain regions, and chronic ethanol treatment did not alter the inhibitory response. These results indicate an apparant lack of adaptation in N-methyl-D-aspartate-stimulated transmitter release following chronic ethanol treatment in this particular paradigm.

Animals↗

PubMed1991

Mechanism of inhibition of N-methyl-D-aspartate-stimulated increases in free intracellular Ca2+ concentration by ethanol.

Dissociated brain cells were isolated from newborn rat pups and loaded with fura-2. These cells were sensitive to low N-methyl-D-aspartate (NMDA) concentrations with EC50 values for NMDA-induced intracellular Ca2+ concentration ([Ca2+]i) increases of approximately 7-16 microM measured in the absence of Mg2+. NMDA-stimulated [Ca2+]i increases could be observed in buffer with Mg2+ when the cells were predepolarized with 15 mM KCl prior to NMDA addition. Under these predepolarized conditions, 100 mM ethanol inhibited 25 microM NMDA responses by approximately 50%, which was similar to the ethanol inhibition observed in buffer without added Mg2+. Ethanol did not alter [Ca2+]i prior to NMDA addition. In the absence of Mg2+, 50 and 100 mM ethanol did not significantly alter the EC50 value for NMDA, but did inhibit NMDA-induced increases in [Ca2+]i in a concentration-dependent manner at 4, 16, 64, and 256 microM NMDA. Whereas NMDA-induced increases in [Ca2+]i were dependent on extracellular Ca2+ and were inhibited by Mg2+, the ability of 100 mM ethanol to inhibit 25 microM NMDA responses was independent of the external Ca2+ or Mg2+ concentrations. Glycine (1, 10, and 100 microM) enhanced 25 microM NMDA-induced increases in [Ca2+]i by approximately 50%. Glycine (1-100 microM) prevented the 100 mM ethanol inhibition of NMDA-stimulated [Ca2+]i observed in the absence of exogenous glycine. MK-801 (25-400 nM) inhibited 25 microM NMDA-stimulated rises in [Ca2+]i in a concentration-dependent manner.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗