Effects-of fluoxetine on sensorimotor and spatial learning deficits following focal cerebral ischemia in rats

Purpose: The present study investigated the effects of fluoxetine, a serotonin reuptake blocker, on behavioral deficits of rats subjected to transient focal cerebral ischemia. Methods: The right middle cerebral artery of rats was occluded for 120 min using the intraluminal filament method. Fluoxetine treatment (5 mg/kg, i.p.) was started 2 days after ischemia induction and treatment was continued for 10 days thereafter. Sensorimotor recovery was assessed using the limb-placing test and cognitive impairment was assessed using a water-maze test at the end of the experiment. Results: Fluoxetine treatment did not improve performance of ischemic rats in the limb-placing test. Nor was the ischemia-induced deficit in the water-maze test affected by fluoxetine. The infarct volumes in the cortex or striatum, determined after the experiment, were not different between ischemic groups. Conclusion: These results suggest that subchronic fluoxetine treatment following experimental focal cerebral ischemia is not detrimental to behavioral outcome, but it also does not enhance spontaneous sensorimotor recovery or attenuate spatial learning deficits.     

Exogenous brain-derived neurotrophic factor prevents postischemic downregulation of [3H]muscimol binding to GABA(A) receptors in the cortical penumbra

We have previously shown that exogenous application of brain-derived neurotrophic factor (BDNF) reduces infarct volume in the cortical ischemic penumbra after experimental focal ischemia [Stroke 31 (2000) 2212-2217]. Since BDNF is known to modulate the expression and function of various neurotransmitter receptors, we addressed the question whether BDNF may act via modification of postischemic ligand binding to excitatory NMDA and AMPA and/or inhibitory GABA(A) receptors, respectively. Transient focal cerebral ischemia was induced in male Wistar rats for 2 h using the suture occlusion technique. A period of 30 min after occlusion of the middle cerebral artery, BDNF (300 microg/kg per hour in vehicle; n=5) or vehicle alone (n=5) was continuously infused intravenously for 3 h. Using quantitative receptor autoradiography, postischemic ligand binding of [(3)H]MK-801, [(3)H]AMPA and [(3)H]muscimol was analyzed in the ischemic core, the ischemic cortical penumbra and corresponding regions of the contralateral hemisphere. Transient focal ischemia caused a significant reduction of [(3)H]muscimol binding to GABA(A) receptors within the ischemic cortical penumbra of placebo-treated rats. This was largely prevented by exogenous application of BDNF. [(3)H]MK-801 and [(3)H]AMPA binding values were also reduced in the cortical penumbra and the corresponding area of the contralateral hemisphere. Our data suggest that the neuroprotective effect of BDNF against ischemic damage in the cortical penumbra may be mediated in part by maintained activity of the inhibitory GABAergic system which likely counteracts glutamate induced excitotoxicity.     

Neuroprotective cannabinoid receptor antagonist SR141716A prevents downregulation of excitotoxic NMDA receptors in the ischemic penumbra

Whether cannabinoids act as neuroprotectants or, on the contrary, even worsen neuronal damage after cerebral ischemia is currently under discussion. We have previously shown that treatment with the cannabinoid (CB1) receptor antagonist SR141716A reduces infarct volume by ∼40% after experimental stroke. Since it is suggested that SR141716A may exert neuroprotection besides its cannabinoid receptor-blocking effect, we addressed the question whether SR141716A may act via modulation of postischemic ligand binding to excitatory NMDA and/or α-amino-3-hydroxy-5-methyl-4-isoxazole-proprionic acid (AMPA) receptors. For this purpose, rats (n = 12) were treated with either intravenous saline (control) or CB1 receptor antagonist SR141716A (1 mg/kg) 30 min after permanent middle cerebral artery occlusion. Five hours after ischemia, quantitative receptor autoradiography was performed using [³H]CP 55,940, [³H]MK-801, and [³H]AMPA for labeling of CB1, NMDA, and AMPA receptors, respectively. Ligand binding was analyzed within the infarct core, cortical penumbra, and corresponding areas of the contralateral hemisphere and compared to that of sham-operated rats (n = 5). Both in ischemic controls and SR141716A-treated rats [³H]CP 55,940 ligand binding was not specifically regulated in the cortical penumbra or contralateral cortex. Importantly, reduced infarct volumes in SR141716A-treated rats were associated with maintained [³H]MK-801 binding to excitotoxic NMDA receptors in the penumbra, compared to a decrease in the control group. In summary, our data suggest that SR141716A may possess additional intrinsic neuroprotective properties independent of receptor-coupled pathways or due to action as a partial agonist.     

Glutamate receptor binding in the frontal cortex and dorsal striatum of aged rats with impaired attentional set-shifting

Aged Long-Evans rats exhibit deficits in attentional set shifting, an aspect of executive function, relative to adult rats. Impairments in set shifting and spatial learning are uncorrelated in aged rats, indicating a possible dissociation of the effects of ageing in prefrontal versus hippocampal systems. Ionotropic glutamate receptor binding was assessed using an in vitro autoradiography method in young and aged rats. The rats had been tested on a set-shifting task that measured attentional set shifts and reversal learning, as well as in a spatial learning task in the Morris water maze. [3H]Kainate, [3H]AMPA and NMDA-displaceable [3H]glutamate receptor binding were quantified in orbital cortex, cingulate cortex, medial frontal cortex, dorsolateral and dorsomedial striatum. Age-related decreases in [3H]kainate binding were apparent in all regions measured. Similarly, NMDA-displaceable [3H]glutamate binding was decreased in the aged rats in all the regions measured except for the medial frontal area where no age effects were observed. [3H]AMPA receptor binding was preserved with age in all the regions measured. Lower levels of [3H]kainate binding in the cingulate cortex were significantly correlated with poorer set-shifting performance, whereas higher levels of NMDA binding in the dorsomedial striatum were correlated with poorer set-shifting performance. There were no significant correlations between the levels of ionotropic glutamate receptors and performance in the reversal task or spatial learning in the Morris water maze. These results indicate that age-related behavioural deficits in attentional set shifting are selectively associated with neurobiological alterations in the cingulate cortex and dorsomedial striatum.     

Regional and selective effects of oestradiol and progesterone on NMDA and AMPA receptors in the rat brain

We investigated the effect of 10 months ovariectomy and a correction therapy, 2 weeks before the rats were killed, of oestradiol, progesterone or their combination on NMDA and AMPA receptor binding in the hippocampus, dentate gyrus, striatum, nucleus accumbens and frontal cortex of the rat brain as well as on amino acid levels in frontal cortex. NMDA and AMPA binding densities were assayed by autoradiography using, respectively, L-[3H]glutamate and [3H]AMPA; amino acid concentrations were measured by high performance liquid chromatograhy (HPLC) coupled with UV detection. Ovariectomy was without effect on NMDA and AMPA binding density in all brain regions assayed except in the hippocampal CA1 region and dentate gyrus where it decreased NMDA binding density compared to intact rats values. Oestradiol restored and increased NMDA binding density in the CA1 subfield and the dentate gyrus of ovariectomized rats but, by contrast, it decreased binding density in the striatum and in the frontal cortex while having no effect in the CA2/3 subfield of the hippocampus and in the nucleus accumbens. Oestradiol was without effect on AMPA binding density in the hippocampus and the dentate gyrus but it reduced AMPA binding density in the striatum, the frontal cortex and the nucleus accumbens. Progesterone, and oestradiol combined with progesterone, decreased NMDA but not AMPA binding density in the frontal cortex of ovariectomized rats, and they were without effect on these receptors in the other brain regions assayed. Amino acid concentrations in the frontal cortex were unchanged after ovariectomy or steroid treatments. The effect of oestradiol in the hippocampus confirmed in the present study and our novel findings in the frontal cortex, striatum and nucleus accumbens may have functional significance for schizophrenia and neurodegenerative diseases.     

An autoradiographic study of [3H]AMPA receptor binding and in situ hybridization of AMPA sensitive glutamate receptor A (GluR-A) subunits following morphine withdrawal in the rat brain

Chronic treatment with opioids is well known to result in the development of physical dependence. More recently, glutamatergic mechanisms have been implicated in expression of the withdrawal syndrome from opioids. To better examine glutamatergic involvement, an autoradiographic study of [3H]AMPA receptor binding and an assessment of in situ hybridization of AMPA sensitive glutamate receptor A (GluR-A) subunits in the rat brain were each performed 7 h after withdrawal from morphine infusion. Animals were rendered dependent by intracerebroventricular (i.c.v.) infusion of morphine (26 nmol/microl/h) via osmotic minipumps for 3 days. Brain sections of 14-microm thickness were incubated with 15 nM [3H]AMPA for quantitation of binding to the AMPA receptor. The probe for in situ hybridization was labeled at its 3' end using terminal deoxynucleotidyl transferase and [35S]dATP. The highest degree of [3H]AMPA binding was shown in the hippocampus. The extent of [3H]AMPA binding was increased significantly in the cortex areas (18-21%), caudate-putamen (20%), and hippocampus (7-9%) of rats following withdrawal from morphine. The highest levels of mRNA for GluR-A, flop and flip subunits, were found in the dentate gyrus and in the CA3 region of the hippocampus, respectively. The levels of mRNA for the flop form of GluR-A were decreased in the CA3 of hippocampus (8%) of the rat brain. The levels of mRNA for the flip form of GluR-A were increased in the parietal cortex (7%) and the entorhinal cortex (8%). Increases in the binding of [3H]AMPA to its receptor may play an important role during withdrawal from morphine dependence.     

Postnatal changes in AMPA receptor regulation by phospholipase A2 treatment of synaptic membranes: temporally differential effects on agonist and antagonist binding.

Previous results have indicated that phospholipase A2 (PLA2) treatment of telencephalic membranes produced opposite effects on [3H]amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) binding in neonatal and adult rats. In the present study, we compared the effects of PLA2 treatment of telencephalic membranes on the binding characteristics of agonists and antagonists of the AMPA receptors in the developing rat brain. Whereas PLA2 treatment of telencephalic membranes from postnatal day (PND) 5 and 10 animals produced an important decrease in [3H]AMPA binding, the same treatment performed on PND 20, 25 and adult membranes resulted in a marked increase in [3H]AMPA binding; the shift from decreased to increased [3H]AMPA binding occurred at about PND 15. In contrast to [3H]AMPA binding, [3H]6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) binding was substantially reduced following PLA2 treatment at PND 5, 10 and 20, and effect due to a decrease in the maximal number of [3H]CNQX binding sites. In adult membranes, the effect of PLA2 treatment on [3H]CNQX binding was markedly reduced when compared to neonatal membranes. Pretreatment of synaptic membranes with PCMBS (a sulfhydryl reagent) increased [3H]AMPA binding in both young (PND 10) and adult telencephalic membranes, without significantly changing [3H]CNQX binding. The various effects of PLA2 treatment on agonist and antagonist binding did not appear to be due to major differences in the pharmacological properties of the AMPA receptors at different ages. The present results indicate that the characteristics of the binding sites for agonists and antagonists of the AMPA receptors are differentially modulated by the lipid environment during the postnatal period.(ABSTRACT TRUNCATED AT 250 WORDS)     

An autoradiographic study of [H]AMPA receptor binding and in situ hybridization of AMPA sensitive glutamate receptor A (GluR-A) subunits following morphine withdrawal in the rat brain

Chronic treatment with opioids is well known to result in the development of physical dependence. More recently, glutamatergic mechanisms have been implicated in expression of the withdrawal syndrome from opioids. To better examine glutamatergic involvement, an autoradiographic study of [³H]AMPA receptor binding and an assessment of in situ hybridization of AMPA sensitive glutamate receptor A (GluR-A) subunits in the rat brain were each performed 7 h after withdrawal from morphine infusion. Animals were rendered dependent by intracerebroventricular (i.c.v.) infusion of morphine (26 nmol/μl/h) via osmotic minipumps for 3 days. Brain sections of 14-μm thickness were incubated with 15 nM [³H]AMPA for quantitation of binding to the AMPA receptor. The probe for in situ hybridization was labeled at its 3′ end using terminal deoxynucleotidyl transferase and [³⁵S]dATP. The highest degree of [³H]AMPA binding was shown in the hippocampus. The extent of [³H]AMPA binding was increased significantly in the cortex areas (18–21%), caudate-putamen (20%), and hippocampus (7–9%) of rats following withdrawal from morphine. The highest levels of mRNA for GluR-A, flop and flip subunits, were found in the dentate gyrus and in the CA3 region of the hippocampus, respectively. The levels of mRNA for the flop form of GluR-A were decreased in the CA3 of hippocampus (8%) of the rat brain. The levels of mRNA for the flip form of GluR-A were increased in the parietal cortex (7%) and the entorhinal cortex (8%). Increases in the binding of [³H]AMPA to its receptor may play an important role during withdrawal from morphine dependence.     

Development of MK-801, kainate,AMPA, and muscimol binding sites and the effect of dark rearing in rat visual cortex

We used quantitative autoradiography to determine whether the development of glutamate receptors correlates with the plastic period for monocular deprivation in rat visual cortex. To study glutamate receptors, we incubated sections of rat visual cortex with tritiated (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]-cyclohepten-5,10imine maleate (MK-801), tritiated kainate, and tritiated amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA). [³H]MK-801 is a noncompetitive ligand for the N-methyl-D-aspartate (NMDA) receptor. [³H]kainate and [³H]AMPA are competitive ligands for non-NMDA receptors. To compare glutamate binding sites with a nonglutamate binding site, we studied [³H]muscimol, which binds to γ-aminobutyric acid (GABA)_A receptors. [³H]MK-801 binding was maximal at postnatal day 26 (P26) and decreased in adulthood. [³H]AMPA binding was maximal at P18. [³H]kainate binding and [³H]muscimol binding were not age dependent. Dark rearing partially prevented the age-dependent decrease in [³H]MK-801 binding but had no effect on [³H]kainate or [³H]AMPA binding. Dark rearing decreased muscimol binding in adult animals. These results suggest that NMDA receptors, but not other glutamate receptors or GABA_A receptors, are likely to be critical for developmental plasticity in rat visual cortex. J. Comp. Neurol. 383:73–81, 1997. © 1997 Wiley-Liss, Inc.     

Chronic alterations in rat brain α-adrenoceptors following traumatic brain injury

Norepinephrine (NE) has been implicated in cerebral plasticity and recovery of function after brain injury. To examine the status of noradrenergic mechanisms in the brain following traumatic brain injury (TBI), male Sprague-Dawley rats underwent right sensorimotor cortex contusions and were observed for the next 30 days for recovery of motor function by measurement of the time taken to perform a modified beam walking task! At 30 days, their brains were assayed by receptor autoradiography for αr- and α2-adrenoceptor binding with 1 nM [3H]prazosin and 1 nM [3H]paraminoclonidine, respectively. One day after contusion, TBI rats took 60% longer to run the beam than sham-lesioned controls. Run times were directly proportional (r = 0.784; P = 0.012) to lesion volume determined at 30 days. The motor deficit persisted for 8 days, after which TBI and control rats had similar run times, largely due to increased run times in sham rats. At 30 days, TBI rats had a generalized, bilateral decrease in [3H]prazosin binding across all brain areas read (F[l,13] = 9.23; P = 0.009) with specific 12%-21% decreases in the cortex contralateral to the lesion and bilaterally in the dorsomedial hypothalamic and three thalamic nuclei. On the other hand, [3H]paraminoclonidine binding did not differ from sham lesion controls in any brain area of TBI rats. Thus, unilateral TBI is followed by widespread, bilateral changes in α1-adrenoceptor binding which would leave the animal vulnerable to any factors which reduced the access of NE to its postsynaptic adrenoceptors. This is compatible with the observation that α1-antagonists and α2-agonists can transiently reinstate the motor deficit after recovery has occurred.     

Postnatal development of the excitatory amino acid system in visual cortex of the rat. Changes in ligand binding to NMDA, quisqualate and kainate receptors

The postnatal development of the ligand binding to N-methyl-D-aspartate (NMDA), quisqualate and kainate receptor sites was examined in whole homogenates of the visual cortex of rats, aged 2-360 days. As selective ligands, [3H]CPP (3-(2-carboxypyperazine-4-yl)-propyl-1-phosphonic acid, [3H]AMPA (RS-alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid) and [3H]KA (kainic acid) were used, respectively. The binding of CPP was low in newborns, rapidly increased from the second postnatal week, reached its maximum between weeks 2 and 3, then slowly declined up to the age of 1 year. In contrast, the binding of AMPA and kainate was high perinatally, increased rapidly up to day 6 after birth to reach an early maximum value, then gradually decreased to adult values which were attained at an age of 3-4 weeks. These age-related changes were derived from alterations in the density of binding sites, which, in the case of AMPA, was accompanied by an increase in binding affinity. The results, compared with the developmental time-course of excitatory synapses, indicate that, in the immature cerebral cortex, NMDA receptors may be primarily involved in synaptic transmission, whereas quisqualate and kainate receptors may play some other (e.g. trophic) roles.     

Discrepancy between cell injury and benzodiazepine receptor binding after transient middle cerebral artery occlusion in rats

We investigated postischemic alterations in benzodiazepine receptor, D1 dopamine receptor, and muscarinic acetylcholine receptor binding after transient middle cerebral artery (MCA) occlusion in rats using [3H]-flumazenil, [3H]-SCH23390, and [3H]-N-methyl-4-piperidyl benzilate ([3H]-NMPB), respectively, as radioligand. These ligand bindings were determined at 3 and 24 h and at 3 and 7 days after ischemia/reperfusion of MCA by using autoradiographic methods. Ischemic cell injury was clearly detected from 3 h after ischemia/reperfusion and progressively increased from 3-24 h after ischemia/reperfusion of MCA. The area of cell injury reached maximum at 24 h after ischemia/reperfusion of MCA. [3H]-SCH23390 binding was reduced to 47% of the contralateral side at 3 days after ischemia/reperfusion of MCA. After 7 days, [3H]-SCH23390 binding was further reduced by 20% in the striatum. [3H]-NMPB binding was slightly decreased in both the striatum and cerebral cortex at 3 days after ischemia/reperfusion of MCA, and [3H]-NMPB binding in the striatum and cerebral cortex were reduced to 42 and 62% of the contralateral side at 7 days after ischemia/reperfusion of MCA. [3H]-NMPB was also decreased at 24 h. In contrast, [3H]-flumazenil binding was not decreased in the striatum and cerebral cortex within 7 days after ischemia/reperfusion of MCA. These results suggest that [3H]-SCH23390 and [3H]-NMPB binding do not correlate with cell injury by ischemia/reperfusion, although vulnerability to ischemia/reperfusion was observed with these receptors. In addition, central benzodiazepine receptor imaging might be essentially stable to neuronal cell injury induced by transient focal cerebral ischemia in rats, in contrast to the results of PET studies.     

Opposite effects of phospholipase A2 on [3H]AMPA binding in adult and neonatal membranes.

In the present study, we compared the effect of phospholipase A2 (PLA2) treatment of synaptic membranes from adult and neonatal rats on the characteristics of [3H]AMPA binding sites. Whereas PLA2 treatment of membranes from adult rats produces an increased affinity for [3H]AMPA binding, the same treatment in neonatal rats results in a decrease in the maximal number of binding sites. Since activation of PLA2 has been proposed to play a critical role in the formation of long-term potentiation (LTP), possibly mediated through a modification of the AMPA receptors, the results strengthen the hypothesis that PLA2-induced modification of [3H]AMPA binding sites is an important component of synaptic plasticity.     

Selective increase of AMPA binding to the AMPA/quisqualate receptor in the hippocampus in response to acute stress.

The binding properties of ligands specific for two subclasses of glutamate receptors were studied by quantitative autoradiography after one hour of acute immobilization/shock stress. [3H]N-(1-(2-thienyl)cyclohexyl)-3,4-piperidine (TCP) and [3H]alpha-amino-3-methylisoxazole-4-propionic acid (AMPA) were used to visualize the N-methyl-D-aspartate receptor and the AMPA/quisqualate receptor types, respectively. While no change was observed in the binding properties of the [3H]TCP, [3H]AMPA binding was significantly increased in several areas of the hippocampus of acutely stressed rats relative to naive controls.     

3H]AMPA binding to glutamate receptor subpopulations in rat brain

The glutamate analog (RS)-alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA), displaced 11% of the binding of L-[3H]glutamate to rat brain membranes, amounting to 22% of the specific binding displaceable by excess non-radioactive glutamate. AMPA-sensitive L-[3H]glutamate binding was additive with that displaced by kainic acid (1 microM) plus N-methyl-D-aspartate (10 microM) when low concentrations of non-radioactive AMPA (1 microM) were employed to determine non-specific background, but partially overlapped when higher concentration of AMPA (100 microM) were used. [3H]AMPA binding was 21% specific (displaceable by non-radioactive 0.1 mM AMPA) in sodium-, calcium- and chloride-free buffer, but increased to over 30% in the presence of 0.1 M chloride. AMPA-sensitive glutamate binding and AMPA binding were both stimulated dramatically by thiocyanate and by several other anions. [3H]AMPA binding activity was resistant to freezing and thawing, optimal at 0-4 degrees C, and detectable at slightly reduced levels by filtration assays and in tissue section autoradiography. AMPA showed a heterogeneous affinity in displacement of L-[3H]glutamate, and [3H]AMPA binding showed heterogeneity with respect to AMPA, quisqualate, and glutamic acid diethyl ester. Scatchard plots gave a best fit for two sites with Kd values of 28 and 500 nM and Bmax values of 200 and 1800 fmol/mg protein, respectively. [3H]AMPA was inhibited by quisqualate (IC50 = 60 nM), L-glutamate (2 microM), (RS)-3-hydroxy-4,5,6,7-tetrahydroisoxazolo-[5,4-c]-pyridine-7-carboxylic acid (7-HPCA, 5 microM), kainic acid (20 microM) and glutamic acid diethyl ester (21 microM) but insensitive to L-aspartate, ibotenic acid, N-methyl-D-aspartate, (RS)-2-amino-phosphonobutyric acid and (RS)-2-amino-phosphonovaleric acid. This is consistent with labeling of a quisqualate-specific subpopulation of glutamate receptors. The high affinity (28 nM) and intermediate affinity (0.5 microM) AMPA sites had similar pharmacological specificity and brain regional distribution as determined by autoradiography. The latter revealed high densities of [3H]AMPA binding in the superficial layers of the cerebral cortex; stratum pyramidale, stratum radiatum, and stratum oriens of the hippocampus; and stratum moleculare of the dentate gyrus. Within the cerebellum, higher densities of binding were observed in the molecular layer than in the granule cell layer. In many regions, [3H]AMPA binding had a similar distribution to that of L-[3H]glutamate binding displaced by AMPA (1 microM).(ABSTRACT TRUNCATED AT 400 WORDS)     

Selective estrogen receptor-alpha but not -beta agonist treatment modulates brain alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors

Estradiol was previously reported to decrease brain alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA)-receptor-specific binding. The contributions of estrogen receptor subtypes in the estradiol modulation of AMPA receptors and its predominant subunit GluR2 are unknown. These experiments investigated whether an estrogenic receptor subtype is involved in the estradiol effect on AMPA-receptor-specific binding and GluR2 mRNA levels. Ovariectomized Sprague-Dawley rats were treated 2 days after ovariectomy for 2 weeks with 17beta-estradiol, an agonist for ERalpha 4,4',4'-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol (PPT), or an agonist for ERbeta 2,3-bis(4-hydroxyphenyl)-propionitrile (DPN) and compared with intact control rats. Uterus weights, used as aperipheral measure of estrogenic activity, were decreased after ovariectomy and increased by estradiol and PPT but not DPN treatments. In prefrontal and cingulate cortices, the striatum, and the nucleus accumbens, ovariectomy increased [3H]AMPA-specific binding compared with intact controls, which was corrected by estradiol treatment. In all these brain regions, PPT, but not DPN, mimicked the estradiol decrease of AMPA-receptor-specific binding; in the cingulate cortex, the effect of PPT did not reach statistical significance. GluR2 mRNA levels of vehicle-treated ovariectomized rats remained unchanged compared with intact rats in the brain regions investigated. Estradiol and PPT treatment but not DPN decreased GluR2 subunit mRNA levels in the prefrontal cortex and the striatum of ovariectomized rats, whereas no significant change was observed in the cingulate cortex or the nucleus accumbens. The present results suggest that an ERalpha is involved in the estradiol modulation of AMPA receptors in the cortex, striatum, and nucleus accumbens.     

Laminar distribution of MK-801, kainate,AMPA, and muscimol binding sites in cat visual cortex: A developmental study

We used quantitative autoradiography to determine whether the development of glutamate receptors correlates with the sensitive period for monocular deprivation in the visual cortex. To study glutamate receptors, we incubated sections of cat visual cortex with tritiated (+)-5-methyl-10, 11-dihydro-5H-dibenzo[a,d]-cyclohepten-5,10imine-maleate (MK-801), tritiated kainate, and tritiated amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA). [³H]MK-801 is a noncompetitive ligand for the N-methyl-D-aspartate (NMDA) receptor. [³H]kainate and [³H]AMPA are competitive ligands for non-NMDA receptors. We used [³H]muscimol, which binds to GABA_A receptors, so that we would have one control ligand that binds to a nonglutamate receptor. When all layers were combined, the results confirmed our previous studies with homogenate binding. [³H]MK-801 and [³H]kainate binding were significantly greater at 42 days than at earlier or later times. [³H]AMPA and [³H]muscimol binding did not show such a peak. This suggests that MK-801 and kainate binding sites are more likely to be involved in plasticity than are AMPA and muscimol binding sites. In layers 2/3, MK-801 had the greatest age-dependent changes; in layers 5 and 6, kainate binding changed most with age. This suggests that the mechanisms of plasticity may vary with cortical layer. © 1996 Wiley-Liss, Inc.     

Non-NMDA glutamate receptor binding in canine brain after global cerebral ischemia and reperfusion

We employed a canine model to test the effects of global cerebral ischemia and reperfusion on binding to α-amino-3-hydroxy-5-methyl-4-isoxazole proprionate (AMPA), kainate (KA), and metabotropic glutamate receptors. Ischemia was induced by 10 min of cardiac arrest, followed by restoration of spontaneous circulation for periods of 0, 0.5, 2, 4, and 24 h. Frozen sections were prepared from parietal and temporal cortex, hippocampus, and striatum, and in vitro autoradiography was performed with one of three radioligands: [³H]AMPA, [³H]KA, or [³H]glutamate (using conditions allowing specific labeling of the metabotropic binding site). In striatum, metabotropic binding was unchanged, whereas AMPA and KA binding decreased by 20–30% at 30 min postischemia, remaining depressed through 24h. In cortex, AMPA and metabotropic binding were decreased at several timepoints after ischemia and recirculation, particularly in parietal cortex, whereas KA binding was unaffected in this tissue. Binding to hippocampal regions was largely unchanged, except for a decrease in KA binding at 2 and 4 h postischemia. These findings contrast with results from parallel studies showing increased striatal binding to NMDA receptors following ischemia. Decreased binding to non-NMDA glutamate receptors in striatum and parietal cortex may serve to protect against damage mediated through these receptors.     

Rehabilitative therapies differentially alter proliferation and survival of glial cell populations in the perilesional zone of cortical infarcts

Rehabilitative therapies after stroke are designed to improve remodeling of neuronal circuits and to promote functional recovery. Only very little is known about the underlying cellular mechanisms. In particular, the effects of rehabilitative training on glial cells, which play an important role in the pathophysiology of cerebral ischemia, are only poorly understood. Here, we examined the effects of rehabilitative therapies on proliferation and survival of distinct glial populations in the perilesional area of photochemically induced focal ischemic infarcts in the forelimb sensorimotor cortex in rats. Immediately after the infarct, one group of animals housed in standard cages received daily sessions of skilled reaching training of the impaired forelimb; a second group was transferred to an enriched environment, whereas a third control group remained in standard cages without further treatment. Functional recovery was assessed in a sensorimotor walking task. To label proliferating cells, bromodeoxyuridine (BrdU) was administered from day 2 until day 6 postinfarct. Proliferation and survival of astrocytes, microglia/macrophages, and immature and mature oligodendrocytes in the perilesional zone were immunocytochemically quantified at day 10 and 42. Using this approach, we demonstrate that enriched environment and reaching training both significantly improve functional recovery of the impaired forelimb. Furthermore, these therapies strongly reduce the proliferation of microglia/macrophages in the perilesional zone, and daily training of the impaired forelimb significantly increased the survival of newly generated astrocytes. Our data, therefore, demonstrate that rehabilitative therapies after cortical infarcts not only improve the functional recovery but also significantly influence the glial response in the perilesional zone.     

Glutamate receptors in striatum and substantia nigra: Effects of medial forebrain bundle lesions

We examined NMDA-sensitive [³H]glutamate, [³H]AMPA, [³H]kainate and metabotropic-sensitive [³H]glutamate binding sites in neostriatum and substantia nigra pars reticulata (SNr) in rats after unilateral 6-hydroxydopamine lesions of the medial forebrain bundle. One week after the lesion, NMDA, AMPA, kainate and metabotropic receptors were decreased in the ipsilateral neostriatum, whereas at three months NMDA receptors were increased while AMPA, kainate and metabotropic receptors were not changed. In the SNr at one week, only AMPA and metabotropic receptors were significantly decreased whereas three months after the lesion NMDA, AMPA and kainate binding sites were decreased. The early decrease of excitatory amino acid receptors in the striatum is likely to reflect degeneration of dopaminergic fibers, suggesting that specific subpopulations of excitatory amino acid binding sites are located on dopaminergic terminals.