Rat Caudatoputamen Research Articles

Stereoselective effects of ketamine on dopamine, serotonin and noradrenaline release and uptake in rat brain slices

Ketamine (2-(2-chlorophenyl)-(1-methylamino)-cyclohexanone) is a rapid-acting dissociative general anaesthetic whose hallucinogenic properties have made it a popular drug of abuse. Ketamine comprises two optical isomers, with differing pharmacology. In the present study, the effects of (+)- and (−)-ketamine on stimulated efflux and reuptake of dopamine (DA), noradrenaline (NA) and serotonin (5-HT) were compared in isolated superfused slices of the rat caudatoputamen (CPu), ventral bed nucleus of the stria terminalis (BSTV) or dorsal raphe nucleus (DRN), respectively. Monoamine efflux was elicited by local electrical stimulation (20 pulses, 100 Hz trains) at tungsten microelectrodes and measured at adjacent carbon fibre microelectrodes using fast cyclic voltammetry (FCV). In CPu (+)-ketamine increased stimulated DA efflux and slowed DA reuptake in a concentration-dependent manner (25–200 μM). At 100 μM (+)-ketamine increased DA efflux by 109±20% (mean±S.E.M., n=13) of control values after 30 min ( P<0.001 versus control) and prolonged uptake half-time ( t 1/2) by 76±38% ( n=9, P<0.001) of control. In contrast (−)-ketamine (100 μM) had no effect on DA efflux or uptake. In DRN, both isomers (100 μM) increased stimulated 5-HT efflux. (−)-Ketamine had a larger effect ( P<0.001), an 88±15% increase in 5-HT efflux ( n=9) versus 46±10% ( n=8) for the (+)-isomer. The isomers had similar effects on 5-HT uptake, increasing t 1/2 by approximately 200%. No evidence of stereospecificity was seen in BSTV: both isomers had small effects (+)- and (−)-ketamine (100 μM) increasing NA efflux by 43±10% ( n=7, P<0.001) and 29±8% ( n=7, P<0.001), respectively. The isomers also had identical effects on NA uptake, each increasing uptake t 1/2 by approximately 100%. In summary, our data show that the optical isomers of ketamine have strikingly different stereospecificity for the monoamine systems and one might predict, therefore, a different psychotomimetic potential.

Strychnine-sensitive glycine receptors in rat caudatoputamen are expressed by cholinergic interneurons

Strychnine-sensitive glycine receptors are ligand-gated anion channels widely expressed in spinal cord and brainstem. Recent functional studies demonstrating glycine-induced release of [ 3H]acetylcholine in rat caudatoputamen suggested the existence of excitatory glycine receptors in that region. Since the expression of glycine receptors in the caudatoputamen had not been reported earlier, we studied the glycine receptor-like immunoreactivity in this structure using a monoclonal antibody (mAb4a) recognizing an epitope common to all of the ligand-binding α-subunit variants of the glycine receptor. [Becker et al. (1993) Brain Res. 11, 327–333; Nicola et al. (1992) Neurosci. Lett. 138, 173–178]. Immunohistochemistry with mAb4a disclosed a specific staining of sparsely distributed large neurons in rat caudatoputamen, displaying an immunoreactive signal of lower intensity than that observed in motoneurons in spinal cord. Fluorescent dual labelling demonstrated that glycine receptor-like immunoreactivity co-localizes with choline acetyltransferase-like immunoreactivity in rat caudatoputamen. All neurons with glycine receptor-like immunoreactivity in the caudatoputamen studied were immunoreactive with choline acetyltransferase, and represented a subpopulation of cholinergic neurons (approximately 90% of the somata with choline acetyltransferase-like immunoreactivity). These results suggest that strychnine-sensitive glycine receptors are present on cholinergic interneurons in rat caudatoputamen, supporting the hypothesis that glycine receptors inducing striatal release of [ 3H]acetylcholine may be localized to cholinergic neurons.

Release and Accumulation of Neurotransmitters in the Rat Brain: Acute Effects of Ethanol In Vitro and Effects of Long‐Term Voluntary Ethanol Intake

Release from and accumulation in tissue slices of some neurotransmitters under acute ethanol in naive rats and in long-term voluntarily ethanol drinking rats were investigated. Slices of the rat caudatoputamen were prelabeled with [3H]choline and release of [3H]acetylcholine was stimulated through either N-methyl-D-aspartate (NMDA) receptors or strychnine-sensitive glycine receptors. Ethanol in vitro at 2 per thousand, 4 per thousand, and 6 per thousand (34 mM, 68 mM, and 102 mM, respectively) concentration-dependently depressed the maximum effect of the concentration-response curve of NMDA in naive rats. In contrast, voluntary ethanol consumption over months led to a significantly enhanced NMDA receptor response characterized by an increase in the maximum effect of the concentration-response curve. The glycine receptor-mediated release of [3H]acetylcholine, which is inhibited by acute ethanol in a competitive-like fashion, was not changed in animals that ingested ethanol over months. Electrically evoked release of [3H]noradrenaline ([3H]NA) and its presynaptic modulation by morphine through mu-opioid receptors in neocortical slices of the rat, preloaded with [3H]NA, was nearly identical in both ethanol-naive rats and in ethanol drinking rats. The accumulation of [3H]gamma-aminobutyric acid in rat cerebellum tissue was neither affected by acute ethanol in vitro nor after chronic ethanol consumption. In summary, long-term voluntary ethanol intake caused a significant increase in NMDA receptor function in the rat caudatoputamen, but did not result in changes in glycine-evoked [3H]acetylcholine release of electrically evoked [3H]NA release modulated by morphine or cerebellar [3H]gamma-aminobutyric acid accumulation.

Distribution of DARPP-32 in the basal ganglia: an electron microscopic study

DARPP-32, a dopamine and cyclic AMP-regulated phosphoprotein, has been studied by light and electron microscopical immunocytochemistry in the rat caudatoputamen, globus pallidus and substantia nigra. In the caudatoputamen, DARPP-32 was present in neurons of the medium-sized spiny type. Immunoreactivity for DARPP-32 was present in dendritic spines, dendrites, perikaryal cytoplasm, most but not all nuclei, axons and a small number of axon terminals. Immunoreactive axon terminals in the caudatoputamen formed symmetrical synapses with immunolabeled dendritic shafts or somata. Neurons having indented nuclei were never immunoreactive. In the globus pallidus and substantia nigra pars reticulata, DARPP-32 was present in myelinated and unmyelinated axons and in axon terminals. The labelled axon terminals in these regions formed symmetrical synaptic contacts on unlabelled dendritic shafts or on unlabelled somata. These data suggest that DARPP-32 is present in striatal neurons of the medium-sized spiny type and that these DARPP-32-immunoreactive neurons form symmetrical synapses on target neurons in the globus pallidus and substantia nigra. The presence of DARPP-32 in these striatal neurons and in their axon terminals suggests that DARPP-32 mediates part of the response of medium-size spiny neurons in the striatum to dopamine D-1 receptor activation.

Effects of somatostatin-14 on the in vitro release of [ 3H]GABA from slices of rat caudatoputamen

Slices (300 μm) of rat caudatoputamen were incubated in Krebs-Henseleit medium and loaded with [ 3H]glutamine, part of which was converted to [ 3H]GABA. This conversion takes place only in GABA-neurons most of which probably contribute to the striatonigral pathway. After a 24 min equilibration period, release of radioactivity was stimulated with veratridine (3.1–4 μmol/l) or K + (15–25 mmol/l) in the absence or presence of somatostatin-14. From the radioactivity released [ 3H]GABA was separated by cationic exchange chromatography and measured. Somatostatin-14 affected the release of [ 3H]GABA in a manner which depended on its concentration as well as on the extent of stimulus-evoked release. Somatostatin-14 (1 nmol/l) enhanced the moderate release (2–4% of tissue content) elicited by veratridine (3.1 μmol/l) or K s+ (20 mmol/l), but had no effect on the more pronounced release (5–8% of tissue content) elicited by veratridine (4 μmol/l) or K + (25 mmol/l). Somatostatin-14 (10 nmol/l) had no effect on the moderate release of [ 3H]GABA, but diminished the pronounced one. Further experiments provided evidence that the somatostatin-14-induced enhancement was not brought about by a direct action on GABA-neurons but was probably indirect, i.e. mediated by other striatal neurons. In contrast, the diminution of the release of [ 3H]GABA caused by somatostatin-14 may be due to its direct action on releasing neurons. Two antisera against somatostatin lowered the pronounced release indicating that endogenous somatostatin may also enhance the release of [ 3H]GABA. In addition, endogenous somatostatin seems also to be able to diminish the release under certain experimental conditions. Somatostatin-14 may be an important modulator of the activity of striatonigral GABA-neurons.

The regulation of cholecystokinin release from rat caudatoputamen in vitro

Cholecystokinin (CCK) is one of the most abundant neuropeptides in the rat caudatoputamen (cp). CCK perikarya innervating the cp are thought to originate in neurons in the claustrum/piriform cortex area of the amygdala. Previous studies on the release of CCK from cp have focused on the influence of dopamine agonists. The present study has examined the influence of other neuroactive substances on the release of CCK. The release of CCK from rat cp slices in vitro stimulated by potassium was quantitated with a specific CCK radioimmunoassay. This potassium-stimulated release of CCK was Ca 2+-dependent. Maximal stimulation of CCK release was observed at 55 mM potassium. Several lines of evidence indicate that the release of CCK from cp is inhibited by some other substance (or substances) released by a Ca 2+-dependent mechanism from cp along with CCK. Release media from cerebral cortex or cp (called ‘conditioned media’ or CM) inhibits the release of CCK from fresh slices of cp but not from cerebral cortex. The release of dopamine from cp is unaffected by CM from cortex or cp. The identity of the substance in CM which inhibits CCK release from cp is still under investigation, though it appears not to be CCK, dopamine, acetylcholine, somatostatin, leucine enkephalin or γ-aminobutyric acid.

Measurement of mRNA specific for preprocholecystokinin in rat caudatoputamen and areas projecting to it

The concentrations of mRNA specific for preprocholecystokinin were measured with a hybridisation procedure after its extraction from rat brain samples. mRNA specific for preprocholecystokinin was not consistently found in the caudatoputamen. In contrast, its concentrations were quite high in the cingulate, subcallosal and parietal cortical areas, which are assumed to send cholecystokinin-containing axons to the caudatoputamen. Also in other areas in which cholecystokinin-pathways to the caudatoputamen may originate, such as the amygdaloid complex, the piriform cortex and the ventral tegmental area/substantia nigra, mRNA specific for preprocholecystokinin could be demonstrated. It is concluded that the caudatoputamen contains no or very few perikarya which synthetize cholecystokinin and that cholecystokinin-peptides which are found in high concentrations in this part of the basal ganglia are located in afferents.

DARPP-32 in the ciliary epithelium of the eye: a neurotransmitter-regulated phosphoprotein of brain localizes to secretory cells.

DARPP-32, a phosphoprotein enriched in dopaminoceptive brain neurons containing the D-1 receptor subtype, probably functions as an intracellular third messenger to mediate some of the physiological effects of dopamine at the D-1 receptor. By immunohistochemistry in rat, cat, Rhesus monkey, and human, we have localized DARPP-32 to the non-pigmented epithelium of the ciliary body, the innermost layer of the bi-layered epithelium responsible for secretion of aqueous humor into the eye. The immunoreactive protein in rat ciliary body, identified by immunolabeling of a ciliary body extract separated by sodium dodecyl sulfate/polyacrylamide gel electrophoresis, is indistinguishable from DARPP-32 derived from rat caudatoputamen. By analogy with brain, we propose that DARPP-32 may act as a third messenger in the ciliary epithelium, probably through a dopaminergic mechanism.

Vasoactive intestinal polypeptide (VIP) inhibits potassium-induced release of cholecystokinin (CCK) from rat caudato-putamen but not from cerebral cortex

CCK release elicited by 40 mM potassium from slices of rat caudato-putamen (cp) was inhibited by VIP. The effect of VIP was maximal at 10 −7 M. VIP does not inhibit CCK release from cerebral cortex at either 10 −7 or 10 −6 M. VIP is known to elevate levels of cAMP in rat brain. VIP inhibition of CCK release appears to be independent of activation of adenylate cyclase because treatment of cp slices with forskolin (2 × 10 −6 to 10 −4 M) does not mimic the inhibitory action of VIP.

Effects of selective dopamine D 2-receptor agonists on the release of cholecystokinin-like immunoreactivity from rat neostriatum

It was investigated whether or not the release of cholecystokinin-like immunoreactivity from slices of rat caudatoputamen could be modulated via dopamine D 2-receptors. The release of immunoreactivity was stimulated by high concentrations of K + (35 to 55 mM). The effects of two D 2-selective dopaminergic agonists, RU 24926 and LY 141865, were tested on the K +-induced release. Both agents enhanced the release of immunoreactivity in a concentration-dependent manner. The effect of RU 24926 was blocked by (−)-sulpiride and domperidone, two selective D 2-antagonists. It is concluded that stimulation of dopamine receptors of the D 2-subtype enhances the release of cholecystokinin-like immunoreactivity from nerve endings in rat neostriatum.

Origin of the cholecystokinin-containing fibers in the rat caudatoputamen.

Large Amounts of cholecystokinin-octapeptide (CCK) are present in the rat caudatoputamen. The peptide occurs in axons and nerve endings but not in perikarya. The origin of CCK in the caudatoputamen was investigated with the use of immunocytochemistry and a radioimmunoassay specific for CCK. Although a small amount of CCK (approximately 30 percent) originates in the amygdaloid complex, the bulk of the peptide (approximately 70 percent) occurs in processes of neurons located ventral to the caudatoputamen, that is, the claustrum or the piriform cortex. The claustrum and piriform cortex receive inputs from various cortical areas and the olfactory system, respectively, and may process information and relay it to the caudatoputamen. Thus CCK may by the transmitter in the final common pathway linking various cortical areas and the olfactory system to the caudatoputamen.

Vascular supply pattern to rat caudatoputamen and globus pallidus: scanning electronmicroscopic study of vascular endocasts of stroke-prone vessels.

The caudatoputamen (CP) and globus pallidus (GP) are supplied by vessels often involved with stroke in both rat and human. The pattern of vascular supply to the CP and GP in rat has, in contrast to humans, been only partially described. The vascular pattern to the rat CP and GP is described utilizing vascular endocasts and scanning electronmicroscopy in aging, normotensive rats. Endocasts were produced by intra-cardiac infusion of Batson's Corrosion Compound. The vascular pattern is complex, involving 1) recurrent vessels from the anterior cerebral artery, 2) branches from the arterial circle rostral or caudal to the origin of the middle cerebral artery (MCA), 3) up to 6 branches from the MCA, and 4) 2 major branches from the caudal part of the arterial circle. The vessels in groups 1--3 were serpentine, their luminal diameters abruptly reduced at branch points, and the angle of departure from the parent vessels approximated 90 degrees. These vessels supplied much of the CP and GP, while group 4 supplied the caudal CP with vessels arranged in a lattice-like fashion from the 2 penetrating parental arteries.