Morphological characteristics and electrophysiological properties of CA1 pyramidal neurons in macaque monkeys.

Little is known about the morphological characteristics and intrinsic electrophysiological properties of individual neurons in the nonhuman primate hippocampus. We have used intracellular recording and biocytin-labeling techniques in the in vitro hippocampal slice preparation to provide quantitative evaluation of the fundamental morphological and intrinsic electrophysiological characteristics of macaque monkey CA1 pyramidal neurons. These neurons have previously been studied in the rat in our laboratory. Monkey CA1 pyramidal neurons have an average soma volume of 3578 microm3, 4.71 basal dendrites with 53 terminal branches for a dendritic length of about 10,164 microm, 1.13 apical dendrites with 47 terminal branches for a dendritic length of about 10,678 microm. In comparison, rat CA1 pyramidal neurons have an average soma volume of 2066 microm3, 3.35 basal dendrites with 29 terminal branches for a dendritic length of about 4,586 microm, 1.43 apical dendrites with 62 terminal branches for a dendritic length of about 8,838 microm. The basic intrinsic electrophysiological properties of CA1 pyramidal cells are similar in monkeys and rats. Monkey CA1 pyramidal neurons have a resting membrane potential of about -62 mV (rat: -62 mV), an input resistance of 35 MOmega (rat: 34-49 MOmega), a rheobase of 0.17 nA (rat: 0.12-0.20 nA) and an action potential amplitude of 83 mV (rat: 71-89 mV). Although morphological differences such as the increased dendritic length may translate into differences in neural processing between primates and rodents, the functional significance of these morphological differences is not yet clear. Quantitative studies of the primate brain are critical in order to extrapolate information derived from rodent studies into better understanding of the normal and pathological function of the human hippocampus.

Facilitated glutamatergic transmission in the striatum of D2 dopamine receptor-deficient mice.

Dopamine (DA) receptors play an important role in the modulation of excitability and the responsiveness of neurons to activation of excitatory amino acid receptors in the striatum. In the present study, we utilized mice with genetic deletion of D2 or D4 DA receptors and their wild-type (WT) controls to examine if the absence of either receptor subtype affects striatal excitatory synaptic activity. Immunocytochemical analysis verified the absence of D2 or D4 protein expression in the striatum of receptor-deficient mutant animals. Sharp electrode current- and whole cell patch voltage-clamp recordings were obtained from slices of receptor-deficient and WT mice. Basic membrane properties were similar in D2 and D4 receptor-deficient mutants and their respective WT controls. In current-clamp recordings in WT animals, very little low-amplitude spontaneous synaptic activity was observed. The frequency of these spontaneous events was increased slightly in D2 receptor-deficient mice. In addition, large-amplitude depolarizations were observed in a subset of neurons from only the D2 receptor-deficient mutants. Bath application of the K+ channel blocker 4-aminopyridine (100 microM) and bicuculline methiodide (10 microM, to block synaptic activity due to activation of GABA(A) receptors) markedly increased spontaneous synaptic activity in receptor-deficient mutants and WTs. Under these conditions, D2 receptor-deficient mice displayed significantly more excitatory synaptic activity than their WT controls, while there was no difference between D4 receptor-deficient mice and their controls. In voltage-clamp recordings, there was an increase in frequency of spontaneous glutamate receptor-mediated inward currents without a change in mean amplitude in D2 receptor-deficient mutants. In WT mice, activation of D2 family receptors with quinpirole decreased spontaneous excitatory events and conversely sulpiride, a D2 receptor antagonist, increased activity. In D2 receptor-deficient mice, sulpiride had very little net effect. Morphologically, a subpopulation of medium-sized spiny neurons from D2 receptor-deficient mice displayed decreased dendritic spines compared with cells from WT mice. These results provide evidence that D2 receptors play an important role in the regulation of glutamate receptor-mediated activity in the corticostriatal or thalamostriatal pathway. These receptors may function as gatekeepers of glutamate release or of its subsequent effects and thus may protect striatal neurons from excessive excitation.

Electrophysiological and morphological analyses of cortical neurons obtained from children with catastrophic epilepsy: dopamine receptor modulation of glutamatergic responses.

The present study examined the electrophysiological effects produced by activation of specific dopamine (DA) receptors and the distribution of DA receptor subtypes and glutamate receptor subunits [N-methyl-D-aspartate (NMDAR1) and GluR1] in cortical tissue samples obtained from children (ages 3 months to 16 years) undergoing epilepsy surgery. DA receptor activation produced differential effects depending on the receptor subtype that was activated. D1 receptor family agonists generally enhanced cortical excitability and favored the emergence of epileptogenic activity. In contrast, D2 receptor family agonists had more variable effects on cortical excitability and the expression of epileptiform discharges. Activation of D1 or D2 receptors decreased the amplitude of non-NMDA-mediated excitatory postsynaptic potentials. In contrast, DA and D1 agonists increased the amplitude of NMDA-mediated potentials. Immunohistochemical analysis showed that the DA receptor subtypes and glutamate receptor subunits examined were present in all cortical layers and areas throughout development. Whole-cell voltage clamp recordings of pyramidal neurons visualized with differential interference contrast optics and infrared videomicroscopy indicated that these neurons displayed a persistent Na(+) current, followed by an outward current. DA reduced the outward current but had little effect on the persistent Na(+) current. These results suggest a dual role for DA's actions in the human cerebral cortex. Activation of D2 receptors or antagonism of D1 receptors may help control seizures in children.

Neonatal hippocampal damage in rats: long-term spatial memory deficits and associations with magnitude of hippocampal damage.

This study investigated the effects of neonatal hippocampal ablation on the development of spatial learning and memory abilities in rats. Newborn rats sustained bilateral electrolytic lesions of the hippocampus or were sham-operated on postnatal day 1 (PN1). At PN20-25, PN50-55, or PN90-95, separate groups of rats were tested in a Morris water maze on a visible "cue" condition (visible platform in a fixed location of the maze), a spatial "place" condition (submerged platform in a fixed location), or a no-contingency "random" condition (submerged platform in a random location). Rats were tested for 6 consecutive days, with 12 acquisition trials and 1 retention (probe) trial per day. During acquisition trials, the rat's latency to escape the maze was recorded. During retention trials (last trial for each day, no escape platform available), the total time the rat spent in the probe quadrant was recorded. Data from rats with hippocampal lesions tested as infants (PN20-25) or as adults (PN50-55 and PN90-95) converged across measures to reveal that 1) spatial (place) memory deficits were evident throughout developmental testing, suggesting that the deficits in spatial memory were long-lasting, if not permanent, and 2) behavioral performance measures under the spatial (place) condition were significantly correlated with total volume of hippocampal tissue damage, and with volume of damage to the right and anterior hippocampal regions. These results support the hypothesis that hippocampal integrity is important for the normal development of spatial learning and memory functions, and show that other brain structures do not assume hippocampal-spatial memory functions when the hippocampus is damaged during the neonatal period (even when testing is not begun until adulthood). Thus, neonatal hippocampal damage in rats may serve as a rodent model for assessing treatment strategies (e.g., pharmacological) relevant to human perinatal brain injury and developmental disabilities within the learning and memory realm.

Agonist-induced morphologic decrease in cellular D1A dopamine receptor staining.

The distribution of D1A dopamine (DA) receptor proteins was assessed by using subtype specific antireceptor antisera after acute DA exposure. The immunofluorescent staining of D1A DA receptor protein expression was examined in (1) stably transfected Chinese hamster ovary (CHO) cells, (2) primary striatal cell cultures, and (3) rat striatal brain slices. After agonist exposure as brief as 2 min and as long as 60 min, profound loss of immunofluorescent D1A receptor protein staining occurred in each paradigm. Additionally in the tissue slice, immunofluorescent neuropil staining for the receptor protein also was attenuated. The DA-induced alteration in receptor protein staining was blocked by the antagonist (+)-butaclamol and by the selective D1-family antagonist SCH 23390. Receptor staining patterns reverted back to the control immunofluorescent distribution within 15 min after removing the agonist from the bath. Immunofluorescence for the second-messenger cyclic AMP increased at all DA exposure times in the three experimental paradigms, was blocked by D1-family antagonists, and decreased to basal staining after brief recovery periods. This demonstrated the functional integrity of the D1A receptor in target cells. Pretreatment with the mitogenic plant lectin concanavalin A blocked the immunofluorescent decrease in receptor staining but not the elevation of the second messenger, indicating a morphologic distinction in these two events, parallel to other biochemical reports. The data suggested that a morphologic basis of acute homologous D1A DA receptor desensitization may be transposition of membrane-surface receptors to a transiently unavailable, intracellular compartment. This finding is supported by specific fluorescence incorporation of FM1-43, used as a marker of endocytosis, in CHO cells treated with DA.

Modulatory actions of dopamine on NMDA receptor-mediated responses are reduced in D1A-deficient mutant mice.

The role of D1 dopamine (DA) receptors in mediating the ability of DA to modulate responses attributable to activation of NMDA receptors was examined in mice lacking D1A dopamine receptors. Specifically, experiments were designed to test the hypothesis that the ability of DA to potentiate responses mediated by activation of NMDA receptors was attributable to activation of D1 receptors. Based on this hypothesis, we would predict that in the D1A mutant mouse, either DA would not induce enhancement of NMDA-mediated responses, or the enhancement would be severely attenuated. The results provided evidence to support the hypothesis. In mutant mice, DA and D1 receptor agonists did not potentiate responses mediated by activation of NMDA receptors. In contrast, in control mice, both DA and D1 receptor agonists markedly potentiated responses mediated by activation of NMDA receptors. The effects of DA in attenuating responses mediated by activation of non-NMDA receptors also were altered in the mutant, suggesting that this action of DA may require coupling or interactions between D1 and D2 receptors. The present studies also provided an opportunity to assess some of the basic electrophysiological and morphological properties of neostriatal neurons in mice lacking D1A DA receptors. Resting membrane potential, action potential parameters, input resistance, excitability, somatic size, dendritic extent, and estimates of spine density in mutants and controls were similar, suggesting that these basic neurophysiological and structural properties have not been changed by the loss of the D1A DA receptor.

Neuromodulatory actions of dopamine on synaptically-evoked neostriatal responses in slices.

The present experiments were designed to further examine the hypothesis that receptor subtype determines the direction of dopamine's (DA) ability to modulate neostriatal neuronal responses. We have reported that DA potentiates responses mediated by activation of N-methyl-d-aspartate (NMDA) receptors, but attenuates responses mediated by activation of non-NMDA receptors in neocortex [Cepeda et al. (1992b) Synapse, 11:330-341] and neostriatum [Cepeda et al. (1993) Proc. Natl. Acad. Sci. U.S.A., 90:9576-9580]. In these studies, responses to excitatory amino acids (EAAs) were evoked by microphoretic application of agonists. The present studies examined whether this differential modulation also applies to components of synaptic responses evoked by electrical stimulation of neostriatal afferents and mediated by activation of specific subtypes of EAA receptors. Using brain slices, the actions of DA and its receptor specific agonists on components of neostriatal synaptic responses that were mediated either by NMDA or non-NMDA receptors were assessed. Responses mediated by NMDA receptors were potentiated by DA while those mediated by non-NMDA receptors were attenuated. These findings provide further support for the hypothesis that the direction of modulatory action of DA is determined by the specific subtype of EAA receptor activated. In addition, the enhancement of NMDA receptor-mediated responses was mimicked by application of SKF 38393, a D1 receptor agonist. Quinpirole, a D2 receptor agonist, consistently attenuated responses mediated by activation of non-NMDA receptors. Thus, the complex modulatory actions of DA are dependent upon combinations of co-activation of specific subtypes of EAA and DA receptors. These findings are of clinical relevance since the actions of DA and EAAs have been implicated in neurological and affective disorders.

Metabotropic glutamate receptor activation selectively limits excitotoxic damage in the intact neostriatum.

The present study was designed to compare the protective consequences of activation of metabotropic glutamate receptors (mGluRs) on N-methyl-D-aspartate (NMDA)- and kainic acid (KA)-induced excitotoxicity in vivo. Pretreatment with the mGluR agonist ISR,3RS-1-aminocyclo-pentane-1,3-dicarboxylic acid (tACPD) limited the anatomical and behavioral consequences of the intrastriatal administration of the NMDA agonist quinolinic acid (QA). In contrast, pretreatment with tACPD did not alter the effects of intrastriatal injection of KA.

Potassium channel blockade does not alter the modulatory effects of dopamine in neostriatal slices.

This study assessed the contribution of K+ conductances to dopamine (DA)-induced modulation of evoked depolarizing synaptic responses (DPSPs) in neostriatal slices obtained from rats. Intracellular recordings of membrane properties and DPSPs evoked by local electrical stimulation were obtained from cells bathed in standard artificial cerebrospinal fluid (ACSF), 5 mM Cs+ in ACSF, 20 mM tetraethylammonium in ACSF, or 1 mM 4-aminopyridine in ACSF. DA altered response amplitude in approximately equal proportions regardless of the presence or absence of these K(+)-channel blockers. These findings suggest that K+ conductances do not provide a major contribution to DA-induced changes in DPSPs in the neostriatum.

Regulation of N-methyl-D-aspartate-induced toxicity in the neostriatum: a role for metabotropic glutamate receptors?

Glutamate release activates multiple receptors that interact with each other and thus determine the response of the cell. Exploring these interactions is critical to developing an understanding of the functional consequences of synaptic transmission. Activation of metabotropic glutamate receptors (mGluRs) inhibits N-methyl-D-aspartate (NMDA)-evoked responses measured electrophysiologically in neostriatal slices. The present study examines the functional consequences of this regulation using infrared differential interference contrast videomicroscopy to measure and characterize glutamate receptor-induced cell swelling in a neostriatal brain slice preparation. This swelling is, in many cases, a prelude to necrotic cell death and the dye trypan blue was used to confirm that swelling can result in the death of neostriatal cells. Activation of mGluRs by the agonist 1-aminocyclopentane-1,3-dicarboxylic acid (tACPD) inhibited NMDA but not amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid/kainate-induced swelling. This regulation was cell-type specific as tACPD did not alter NMDA-induced swelling in pyramidal cells of the hippocampus. Importantly, these findings could be extended to in vivo preparations. Pretreatment with tACPD limited the size of lesions and associated behavioral deficits induced by intrastriatal administration of the NMDA receptor agonist quinolinic acid.

Behavioral correlates of vitamin D deficiency.

Rats deprived of vitamin D at weaning were compared to control rats on open field, stabilimeter, radial arm maze and spatial reversal tasks in order to test the hypothesis that vitamin D deficiency alters behavior and learning. The deficient animals engaged in statistically less open field rearing activity and spent more time each day negotiating the radial maze than did the control rats. These findings are consistent with the known influence of vitamin D on the musculoskeletal system. The deprived rats did not differ from the control animals on the learning measures. This would indicate that vitamin D deficiency may not significantly impair cognitive functions in young adult rats.

Effects of medial frontal cortex lesions on DRL performance in rats.

Rats with medial frontal cortical lesions (MFC) and control animals with sham operations were tested on an ascending series of 8 DRL schedules of reinforcement ranging from 5 to 70 seconds. Rats with MFC lesions did not differ from the control group in performance when response rate, responses per reinforcement and efficiency were analyzed. These results indicate that deficits in "response inhibition" often displayed by rats with MFC lesions may be task-specific.

Conditionability of caudate dopamine activity in the caudate nucleus of cats demonstrated by in vivo electrochemistry.

An in vivo electroanalytic method was used to measure dopamine and dopamine metabolite levels in the anterior caudate of freely behaving cats. Food reinforcement was made contingent on either increasing or decreasing dopamine metabolite levels. The cats were found to be capable of contingent metabolite level variation in accord with reinforcement contingency. The metabolite variations were observed as both short-term and long-term changes, and were significantly different than variations obtained during random reinforcement. No distinctive overt behavioral responses were observed to accompany any of the reinforcement conditions. These results indicate that central dopaminergic activity can be significantly altered through extrinsic control and further imply that specific stimulus environments may contribute to the regulation and/or disregulation of neurochemical brain systems.