bluuebunny/crossref_metadata_2025 huggingface dataset:
Interaction of β‐funaltrexamine with [3H]cycloFOXY binding in rat brain: Further evidence that β‐FNA alkylates the opioid receptor complex
Authors: Rothman Richard B. , Bykov Victor , Mahboubi Artin , Long Joseph B. , Jiang Qi , Porreca Frank , de Costa Brain R. , Jacobson Arthus E. , Rice Kenner C. , Holaday John W.
Published: December 2004

β‐Funaltrexamine (β‐FNA) is an alkylating derivative of naltrexone. In addition to acting as an irreversible inhibitor of μ‐receptor‐mediated physiological effects, intracerebroventricular (i.c.v.) administration of β‐FNA to rat attenuates the ability of selective δ receptor antagonists and naloxne to reverse δ receptor‐mediated effects. Moreover, recent work demonstrated that i.c.v. administration of β‐FNA alters the conformation of the opioid receptor complex, as inferred by a decrease in the Bmax of the lower affinity [3H][D‐ala2, D‐leu5]enkephalin binding site. Consistent with the decreased potency of naloxone as an inhibitor of δ receptor mediated effects, β‐FNA doubled the naloxone IC50 for displacing [3H][D‐ala2, D‐leu5]enkephalin from its lower affinity binding site. These data collectively support the hypothesis that the opioid receptor complex postulated to mediate μ‐δ interactions in vivo is identical to the opioid receptor complex as defined by vitro ligand binding studies. A direct prediction of this hypothesis is that β‐FNA should increase the Kd of antagonists for the μ binding site (μcx) of the receptor complex. The data reported in this paper demonstrate that β‐FNA doubled the IC50 of the potent narcotic antagonist, 6‐desoxy‐6β‐fluoronaltrexone (cycloFOXY) for displacing [3H][D‐ala2, D‐leu5]enkephalin from its lower affinity binding site, and doubled the Kd of [3H]cycloFOXY for its μ binding site, providing additional data that the μ binding site labeled by [3H]cycloFOXY is the μ binding site of the opioid receptor complex. β‐FNA also altered the K binding site labeled by [3H]cycloFOXY, and when administered intrathecally to mice, β‐FNA produced a longlasting antinociception in the acetic acid writhing test.
Organization of mitochondria in olfactory bulb granule cell dendritic spines
Authors: Cameron Heather A. , Kaliszewski Christine K. , Greer Charles A.
Published: December 2004

In contrast to dendritic spines with only postsynaptic functions, the spines of olfactory bulb granule cells subserve both pre‐and postsynatic roles. In single sections these spines were previously seen to contain mitochondria, most likely needed to provide energy for presynaptic functions, but their frequency and distribution were unkown. In order to understand the organization of mitochondria in these specialized dendritic appendages, we have studied the geometry and cytoplasmic organization of granule cell spines with computer‐assisted reconstructions of serial electron micrographs. The spine heads were seen to be elliptical in shape with a single pair of reciprocal synapses on the concave face apposed to the mitral/tufted cell dendrite. Mitochondria were found localized in the spine neck as well as the spine head and often extended between the two compartments, Based or their variable distribution it seems reasonable to suggest that these mitochondria are motile and move in and out of spine compartments from the parent dendrite. Spine apparatus was apparent in most of the spines as membrane bound cisterns of smooth endoplasmic reticulum located close to mitochondria. The possible role of spine apparatus in facilitating the movement of mitochondira in the necks and heads of granule cell spines in the absence of microtubules is discussed.
Comparison of the effects of various typical and atypical antipsychotic drugs on the suppressant action of 2‐methylserotonin on medial prefrontal cortical cells in the rat
Authors: Ashby Charles R. , Minabe Yoshio , Edwards Emmeline , Wang Rex Y.
Published: December 2004

In this study, we report the effects of various typical and atypical antipsychotic drugs (APDs) on the suppressant action of microiontophoretically applied 2‐methylserotonin (2‐Me‐5HT, a 5‐HT3agonist) on medial prefrontal cortical (mPFc) cells. The microiontophoresis of 2‐Me‐5HT (10–80 nA) produced a current‐dependent suppression of mPFc cells' firing, and this effect was blocked by various 5‐HT3 antagonists. The microiontophoresis of the atypical APDs clozapine and a structurally related compound, RMI 81, 582, mimicked the action of the 5‐HT3 antagonists. In addition, the intravenous administration of clozapine and RMI 81, 582 antagonized the suppressant action produced by the iontophoretic application of 2‐Me‐5HT on mPFc cells. However, the suppressant action of 2‐Me‐5HT was not blocked by the typical APDs haloperidol and chlorpromazine. The putative atypical APDs risperidone, setoperone, CL 77328, SCH 23390, CGS 10746B, 1‐sulpiride, and thioridazine were ineffective in antagonizing 2‐Me‐5HT's action. Overall, our results suggest that the majority of putative atypical APDs do not interact with 5‐HT3 binding sites in the brain. Whether the interaction of clozapine and RMI 81, 582 with 5‐HT3 sites is correlated with their therapeutic efficacy or lower potential to induce neurological side effects remains to be determined.
Ultrastructure of aberrant serotonin‐immunoreactive fibers in the caudate putamen complex of the aged rat
Authors: van Luijtelaar Mariëlle G. P. A. , Wouterlood Floris G. , Tonnaer Jeroen A. D. M. , Steinbusch Harry W. M.
Published: December 2004

Degeneration of neurons in the central nervous system is associated with morphological changes. Previous observations made at the light microscopical level indicated degeneration of serotonin‐immunoreactive (IR) fibers in the aged rat brain. In this study, a comparison at the ultrastructural level was made between serotonin‐IR normal thin and aberrant swollen varicose fibers in the caudate‐putamen complex of the aged rat. Ultrastructural features such as the size and content of the thin varicose fibers resembled those in the caudate‐putamen complex of the young rat as reported by others. The aberrant profiles were swollen, reaching a size of 6 μm. Their vesicles varied in size and were no longer uniformly round. Moreover, distorted mitochondria and membranefilled vacuolelike structures were a common feature of the aberrant profiles. These changes are indicative of a degenerative process and give further evidence that, whereas many serotonergic fibers are preserved at high age, other serotonergic fibers are degenerating in the caudate‐putamen complex of the aged rat.
Characterization and distribution of [3H]ohmefentanyl binding sites in the human brain
Authors: Wang Hong , Sarrieau Alain , Pélaprat Didier , Roques Bernard P. , Vanhove Alain , Kopp Nicolas , Chi Zhi‐Qiang , Rosténe William
Published: December 2004

Binding properties and localization of [3H]ohmefentanyl, a new ligand for μ opioid receptors, were investigated on normal human brain sections. Binding assays performed at the level of the basal ganglia revealed: (1) a steady‐state binding reached after 60 min incubation at room temperature, (2) the presence, in saturation experiments, of an apparent single class of binding sites with a Kd = 1.68 ± 0.45 nM and a Bmax = 162 ± 9 fmol/mg protein, (3) an order of potency to inhibit [3H]ohmefentanyl binding as follows: ohmefentanyl > [D‐Ala2, MePhe4, Gly‐ol5]enkephalin (DAGO) > ethylketocyclazocine (EKC) ≫ Tyr‐D‐Ser(OtBu)‐Gly‐Phe‐Leu‐Thr(OtBu) (BUBU) and U‐50, 488H. Quantitative autoradiography showed an heterogeneous distribution of [3H]ohmefentanyl binding sites with the highest densities in amygdala, medial geniculate body, thalamus, and caudate nucleus. Binding characteristics and anatomical distribution also show that [3H]ohmefentanyl may bind to a small proportion of additional sites called “DAGO‐inaccessible [3H]ohmefentanyl specific binding sites.” [3H]Ohmefentanyl binding to these sites can be partly inhibited by s̀ ligands such as 1, 3‐di‐o‐tolyguanidine (DTG) and haloperidol. However, unlabeled DAGO inhibited more than 80% of [3H]ohmefentanyl specific binding in most of the human brain regions studied, suggesting that the major population of sites labeled by [3H]ohmefentanyl represented μ opioid receptors.
Clinical and biochemical aspects of depressive disorders: I. Introduction, classification, and research techniques
Authors: Caldecott‐Hazard Sally , Guze Barry H. , Kling Mitchel A. , Kling Arthur , Baxter Lewis R.
Published: December 2004

The present review focuses on recent data from clinical and animal research concerning the biochemical bases of depressive disorders, diagnosis, and treatment. In addition to integrating these data, problems and future directions in this research are discussed. The review is presented in three parts. This study, Part I, describes diagnostic classification schemes for depressive disorders, some epidemiological and biological correlates of the classifications, and research techniques for investigating depressive disorders. Research techniques include animal models, human biochemical techniques, and Postron Emission Tomography. In a future issue, Part II will discuss various transmitter/receptor theories of depressive disorders, e.g., noradrenergic, serotonergic, cholinergic, and dopaminergic, GABAergic, and peptidergic theories. Also in a future issue, Part III will discuss treatments for depression and some of the controversies in the field.
Influence of age on N‐methyl‐D‐aspartate antagonist binding sites in the rat brain studied by in vitro autoradiography
Authors: Miyoshi Rie , Kito Shozo , Doudou Naomi , Nomoto Teruko
Published: December 2004

The N‐methyl‐D‐aspartate (NMDA) receptor complex has been considered to consist of an L‐glutamate recognition site, a strychine‐insensitive glycine modulatory site, and a voltage‐dependent cation channel. In this study, an effect of age on NMDA antagonist binding sites was investigated through quantitative in vitro autoradiography with 3H‐3‐((+)‐2‐carboxypiperazin‐4‐yl)propyl‐1‐phosphonic acid (CPP). 3H‐CPP binding sites were most concentrated in the hippocampus and cerebral cortex where NMDA receptors have been demonstrated to be involved in synaptic transmission. In aged rats, 3H‐CPP binding sites in the hippocampus and cerebral cortex were not significantly changed. As for other brain regions, there was an age‐dependent decline of binding sites only in the caudate‐putamen and nucleus accumbens. Our previous study revealed that strychine‐insensitive glycine receptors were markedly reduced in telencephalic regions in the aged rat brain. Taking these findings into consideration, is concluded that glycine receptors but not NMDA antagonist binding sites are severely altered in telencephalic regions of aged animals. It is considered that within the NMDA receptor complex, glycine receptors may be selectively affected in the aging process.
Neuroanatomic specificity and time course of alterations in rat brain serotonergic pathways induced by MDMA (3,4‐methylenedioxymethamphetamine): Assessment using quantitative autoradiography
Authors: Battaglia George , Sharkey John , Kuhar Michael J. , de Souza Errol B.
Published: December 2004

The Widely abused “designer” durg MDMA (3,4‐methylenedioxymethamphetamine) has been shown to caused marked and long‐lasting changes in brain serotonergic systems. The present study uses quantitative in vitro autoradiography of 3H‐paroxetine labeled 5‐HT uptake sites to assess the time‐dependent effects of MDMA on 5‐HT neurons in specific neuroanatomic loci. Following treatment with MDMA (20 mg/kg, b.i.d. for 4 days), marked decreases in 5‐HT uptake sites were observed in a number of brain regions known to receive projections of ‐5HT neurons. These regions included cerebral cortex, caudate nucleus, hippocampus, nucleus accumbens, olfactory tubercle, superior and inferior colliculi, geniculate nuclei, and most thalamic nuclei. In contrast, other areas such as the septal nuclei and some thalamic nuclei which also receive 5‐HT projections were not substantially affected by this drug. In most regions, decreases in 5‐HT uptake sites occurred within 24 hours of the last dose of MDMA and persisted at the 2 week time point. Some regions such as dorsal striatum exhibited a time‐dependent reduction with greater reductions occurring at 2 weeks rather than immediately following the MDMA treatment regimen. The density of 5‐HT uptake sites in other regions such as endopiriform nucleus and substantia nigra at the 2 week versus 18 hour time point indicated some degree of region‐specific recovery. Regions which demonstrated no significant reduction in 5‐HT uptake sites included the dorsal and median raphe nuclei, ventral tegmental area, central grey, interpeduncluar nucleus, locus coerulus, pontine reticular formation and cerebellum. Likewise, region containing 5‐HT axons of passage (e.g., indusium griseum and lateral hypothalamus) appeared to be insensitive to the neurotoxic effects of MDMA on 5‐HT neurons. Furthermore, the neurotoxic effects of MDMA showed specificity in that the catecholamine neurons labeled by 3H‐mazindol were unaffected by the treatment regimen. These data indicate that the preferential degeneration of serotonergic neurons by MDMA is mediated primarily at 5‐HT terminal regions, whereas regions containg 5‐HT perikarya and axons of passage remain relatively unaffected. In addition, the observed time‐dependent reductions and recovery of 5‐HT uptake sites which were detected within 2 weeks of the treatment regimen in certain brain regions suggest region‐specific differences in recovery of 5‐HT systems from MDMA‐induced lesion.
Calcitonin gene‐related peptide in the amygdaloid complex of the rat: Immunohistochemical and quantitative distribution, and drug effects on calcium dependent, potassium‐evoked in vitro release
Authors: Haring C. , Humpel C. , Skofitsch G. , Krobath J. , Javorsky F. , Saria A.
Published: December 2004

The amygdaloid complex is an area with a high concentration of calcitonin gene‐related peptide. In the present paper, immunohistochemical studies revealed a dense innervation of the central nucleus originating most probably from the bed nuclei of the stria terminalis. For determination of tissue concentrations of calcitonin gene‐related peptidelike immunoreactivity, the amygdaloid complex was dissected into four parts. The distribution was found to be uneven with the highest concentration (1153.3 fmol/mg protein) in the portion including the nucleus amygdaloideus centralis. High performance liquid chromatographic analysis from an extract of amygdaloid tissue showed that 97% of the calcitonin gene‐related peptidelike immunoreactivity measured by radioimmunoassay is authentic rat calcitonin gene%related peptide alpha or beta. Release of calcitonin gene‐related peptidelike immunoreactivity was measured in superfused slices of amygdalae pooled from three rats. High potassium (60 mM) caused a significant release of calcitonin gene‐related peptidelike immunoreactivity (from 0.88% of total tissue content to 1.91%) from the amygdaloid complex in vitro, which was blocked in calcium‐free buffer. Pretreatment with haloperidol or clozapine caused a significant reduction of the 60 mM potassium‐evoked release, compared with a saline treated control group (control 21.0 fmol; haloperidol 2.8 fmol; clozapine 8.8 fmol) and an increase of tissue levels after haloperidol treatment by 43%. These results demonstrate that calcitonin gene‐related peptide is integrated in amygdaloid functions and possibly a target for actions of neuroleptic drugs.
Beta‐adrenergic receptor subtypes in the basal ganglia of patients with Huntington's chorea and Parkinson's disease
Authors: Waeber Christian , Rigo Monique , Chinaglia Giorgia , Probst Alphonse , Palacios José M.
Published: December 2004

The density of [125I]iodo‐cyanopindolol binding to beta‐1 and beta‐2 adrenergic receptors was studied in post mortem basal ganglia samples of Huntington's chorea and Parkinson's disease patients using autoradiography. Whereas no significant changes were observed in sections from Parkinson's and Huntington's chorea grade 2 patients, a nearly complete loss of beta‐1 binding sites was observed in the basal ganglia of Huntington patients at later stages of the disease. The concentration of beta‐2 receptors was increased by a factor 2 in the posterior putamen of all choreic cases. These results are consistent with the view that beta‐1 receptors are predominantly located on a subpopulation of neurons which degenerate at late stages of Huntington's chorea, while beta‐2 receptors are present mainly on glial elements.
The effect of the acetylcholine transport blocker vesamicol on central cholinergic pressor neurons
Authors: Buccafusco Jerry J. , Wei Jian , Kraft Katherine L.
Published: December 2004

Vesamicol (AH 5183) inhibits the uptake of acetylcholine into cholinergic neuronal storage vesicles. Earlier in vitro studies have demonstrated that such inhibition can lead to a failure of transmission, particularly in peripheral cholinergic tissues. The present study was designed to determine whether vesamicol could inhibit central cholinergic transmission in conscious freely moving rats. Central (lateral cerebroventricular) injection of 20 μg of vesamicol significantly reduced the hypertensive and bradycardic response to subsequent central injection of physostigmine in spontaneously hypertensive rats. Inhibition of the pressor response was greatest when physostigmine was administered 1 hr after vesamicol. Acetylcholine and choline levels were determined in three brain regions derived from rats treated one hr earlier with either vehicle or vesamicol. Acetylcholine levels were found to be unaltered after vesamicol treatment, although choline levels were significantly higher in two brain regions. These results are consistent with the ability of vesamicol to inhibit the function of central cholinergic cardiovascular regulatory neurons. The mechanism for this inhibition is not related to depletion of total brain acetylcholine content but may be due to depletion of a small critical pool of transmitter.
Actions of phosphomonoesters on CA1 hippocampal neurons as revealed by a combined electrophysiological and nuclear magnetic resonance study
Authors: Bradler James E. , Barrionuevo German , Panchalingam Kanagasabi , McKeag Dennis , Pettegrew Jay W.
Published: December 2004

Phosphomonoesters (PMEs), precursors of membrane phospholipids, are found in high levels in the developing brain and in Alzheimer's disease brain. The present study details the neurophysiological and metabolic effects of acute PME elevation on the Fisher 344 rat in vitro hippocampal slice. Two abundant PMEs, phosphoethanolamine (PE) and L‐phosphoserine (PS), reliably altered properties of synaptic transmission at the Schaffer collateral/commissural—CA1 cell synapse. Specifically, PE reversibly depressed the amplitude of population EPSPs at millimolar concentrations but had no effect at micromolar concentrations. PS had biphasic effects on population EPSPs, inducing first a reduction followed by an enhancement of response amplitude. In contrast to PE, the effects of PS were not reversible; population EPSPs were augmented during the wash of PS, and the CA3 region generated evoked (but not spontaneous) epileptiform discharges. 31P nuclear magnetic resonance spectroscopy revealed enhanced slice uptake of PS compared to PE. There was no significant effect of PE on slice high‐energy phosphates but incubation with PS significantly lowered slice phosphocreatine (PCr) and ATP concentrations. These observations indicate that the slice uptake of PS could be energy requiring and the enhanced response amplitude observed at 5 mM PS also could produce a drain on high‐energy phosphates. Possible modes of PME action on hippocampal physiology are discussed.
Comparative analysis of the effects of iontophoretically applied dopamine in different regions of the rat brain, with special reference to the cingulate cortex
Authors: Beauregard Mario , Ferron André , Descarries Laurent
Published: December 2004

A systematic comparison of the effects of iontophoresed dopamine (DA) was carried out in the neostriatum (NS), nucleus accumbens (Acb) and anterior cingulate (ACg), prefrontal (PF) and parietal (Par) cortex of urethane‐anesthetized rats, before and after treatment with the specific DA uptake blockers GBR 12909 and Bupropion. Similar experiments were also conducted after DA denervation with 6‐hydroxydopamine and after DA depletion with α‐methyl‐p‐tyrosine. The average rate of spontaneous neuronal firing was comparable in all regions, except in the NS after DA depletion. A majority of the units were inhibited by DA in every region and condition tested. As assessed with the IT50 index, the responsiveness to DA was not markedly different between regions, indicating that the postsynaptic sensitivity to this amine is independent of the density of DA receptors and of DA innervation. In contrast, the average duration of DA inhibitions (RT90) was considerably longer (5‐fold) in the intact ACg than in the PF, Par, NS, or Acb. Moreover, treatment with both DA uptake blockers reduced the duration of DA inhibitions in ACg (4‐ to 9‐fold); while lengthening it in PF, NS and Acb; and having no apparent effect in Par. DA depletion and DA denervation also reduced the duration of the DA inhibitions in ACg without effect in Par. Taken together, these results provide further evidence for the existence of a presynaptic, positive‐feedback mechanism in ACg, triggered by DA, and favouring the further release of this transmitter upon its reuptake in DA nerve terminals.
Severe depletion of cocaine recognition sites associated with the dopamine transporter in Parkinson's‐diseased striatum
Authors: Kaufman Marc J. , Madras Bertha K.
Published: December 2004

The cocaine congener [3H]CFT, also designated [3H]WIN 35,428 (2β;‐carbomethoxy‐3β;‐(4‐fluorophenyltropane)), labels cocaine receptors associated with the dopamine transporter in primate striatum. Autoradiogrpahic distribution of [3H]CFT binding (5 nM) in human postmortem control and Parkinson's‐diseased striatal tissue sections was compared. In control tissue, high and comparable levels of [3H]CFT binding were observed in the putamen and caudate nucleus. At least 90–99% of total [3H]CFT bound was inhibited by (—)‐cocaine (30 μM), suggesting that a high proportion of [3H]CFT is specifically bound. In Parkinson's‐diseased tissue, binding sites for [3H]CFT were reduced by 80% in the caudate nucleus and 96% in the putamen. This pattern of depletion parallels the previously reported loss of dopamine in these brain regions (Kish, Shannak, and Hornykiewicz, New Engl. J. Med., 318:876–880, 1988). In the dorsal caudate nucleus of Parkinson's‐diseased tissue, a lateral‐to‐medial gradient of [3H]CFT binding was observed, with the lateral caudate more severely depleted than the medial caudate. The marked decrease of [3H]CFT binding sites in Parkinson's diseased striatum supports the following conclusions: (1) the dopamine transporter is localized primarily on presynaptic nigrostriatal terminals; (2) in the caudate and putamen, cocaine recognition sites are associated primarily with the dopamine transporter; (3) the low level of nonspecific binding of [3H]CFT and the marked depletion of [3H]CFT‐labeled sites suggest that radiolabeled derivatives of CFT or its congeners may be suitable imaging probes for presynaptic dopamine nerve terminals.
Distribution of somatostatin‐immunoreactive cell bodies and fibers in the neocortex of Macaca fuscata
Authors: Iritani Shuji , Satoh Keiji
Published: December 2004

The distribution of somatostatin‐immunoreactive cell bodies and processes was studied in the cerebral cortex of the macaque monkey (Macaca fuscata), by applying an immunohistochemical technique with a monoclonal antibody raised against somatostatin tetradecapeptide. Many somatostatin‐immunoreactive cell bodies and processes were observed in all regions of the cerebral cortex, i.e., frontal, parietal, temporal, insular, occipital, and cingulate cortices, and also in the underlying white matter. Three types of somatostatin‐contaning cell bodies were distinguished in the cerebral cortex. These cell bodies were distributed in layers II to VI of the cortex, and also in the underlying white matter. There were dense deposits of somatostatin‐containing granular structure in layers I and II, and many somatostatin‐containing processes in layers IV and V. The present observations demonstrate that in the primate neocortex somatostatin 14‐containing neuronal systems are as highly developed as other prosomatostatin‐derived peptides but differe from the latter systems in terms of cell morphology and fiber distribution.
Basal extracellular dopamine is decreased in the rat nucleus accumbens during abstinence from chronic cocaine
Authors: Parsons L. H. , Smith A. D. , Justice J. B.
Published: December 2004

Rats were treated for 10 days with cocaine (20 mg/kg, i.p.) followed by either 1 or 10 days of abstinence. On the test day a microdialysis method was performed in which dopamine (DA) was added to the perfusate at concentrations above and below the expected extracellular concentration (0, 2.5, 5, and 10 nM) to generate a series of points that can be interpolated to determine the concentration of no net flux, which represents the extracellular DA concentration. The slope of the line generated by this method is the in vivo recovery of the dialysis probe. After 1 day of abstinence, there was no significant difference in basal DA levels in the nucleus accumbens (N ACC) between cocaine treated (4.1 ± 0.3 nM; mean ± SEM) and saline‐treated (3.9 ± 0.2 nM) groups. However, there was a significant increase in the slope of the cocaine‐treated group (0.91 ± 0.04 vs. 0.67 ± 0.08; P >0.03). After 10 days of abstinence, there were reduced basal extracellular levels of DA in the N ACC of the cocaine‐treated group as compared with saline‐treated controls (P <0.002). The basal extracellular DA concentration in the N ACC was 2.1 ± 0.3 nM for the cocaine group and 3.9 ± 0.2 nM for the control group. The slopes of the curves were not significantly different for the cocaine (0.63 ± 0.07) and saline (0.64 ± 0.09) groups. There was a significant interaction between cocaine and the length of abstinence with respect to the reduction in basal extracellular DA in the N ACC (P <0.0008) and also an interaction between cocaine and the length of abstinence with respect to the slope of the regression lines (P <0.03). The results suggest that both basal levels and the dynamics of extracellular DA in the N ACC are altered by chronic cocaine. These alterations may have relevance to the dopamine depletion hypothesis of cocaine addiction.
Characterization of the distribution of Gαo in rat striatal synaptosomes and its colocalization with tyrosine hydroxylase
Authors: Wolf Marina E. , Granneman James G. , Kapatos Gregory
Published: December 2004

Dopaminergic striatal synaptosomes can be detected and isolated with a fluorescence‐activated cell sorter (FACS). In the present study, two antigens were detected simultaneously with primary antisera raised in different species and species‐specific fluorescent secondary antibodies with different emission spectra. Double‐label FACS analysis was used to determine whether tyrosine hydroxylase (TH) and the alpha subunit of Gαo (Gαo) are colocalized in striatal synaptosomes. Rabbit antibodies generated against a synthetic fragment of Gαo (corresponding to amino acids 22–35) combined with fluorescein‐conjugated secondary antibodies were used to detect Gαo‐containing striatal synaptosomes. Preadsorption of Gαo antiserum with the synthetic peptide antigen reduced labeling to the level obtained with preimmune serum. Approximately 65–75% of striatal synaptosomes were specifically labeled by Gαo antiserum. Tyrosine hydroxylase‐containing synaptosomes were detected with a mouse monoclonal antibody to TH and R‐phycoerythrin‐conjugated secondary antibody. They comprised 15–17% of total striatal synaptosomes. Double‐label studies indicated that at least 50% of TH‐containing synaptosomes also contained Gαo. These finding suggest that Gαo may not be a protein component of all striatal nerve terminals, and provide a basis for a role for Gαo in signal transduction within subpopulations of intrinsic and afferent nerve terminals, including those of nigrostriatal dopamine neurons.
Dissociative anesthesia and striatal neuronal electrophysiology
Authors: Kelland Mark D. , Chiodo Louis A. , Freeman Arthur S.
Published: December 2004

Compared with results obtained in locally anesthetized, paralyzed rats, the dissociative anesthetic ketamine did not alter either the number of spontaneously active striatal neurons or the basal firing rate of striatal neurons; 90% of these cells exhibited the type I striatal neuron waveform. Chloral hydrate anesthesia suppressed both the occurrence and the firing rate of spontaneously active type I cells, but did not alter the activity of type II striatal neurons. Cortical stimulation preferentially activated type II cells in paralyzed rats and in chloral hydrate‐anesthetized rats. Thus, under dissociative anesthesia it is possible to study spontaneously active type I striatal neurons. However, a method of activation such as cortical stimulation is necessary to study type II striatal neurons.
Midbrain dopamine system electrophysiological functioning: A review and new hypothesis
Authors: Bunney Benjamin S. , Chiodo Louis A. , Grace Anthony A.
Published: December 2004

The pathogenesis of a variety of neurological and mental disorders has been attributed to the malfunctioning of central dopaminergic systems. Twenty years of investigation concerning the way that dopamine systems function in the brain has yielded a great deal of information about the control and behavior of these systems. This collection of seemingly disparate facts is reviewed. A hypothesis is then presented that attempts to synthesize from these facts a view of the electrophysiological functioning of midbrain dopamine systems that may be helpful in understanding brain mechanisms underlying the pathology of such disorders as Parkinson's disease and schizophrenia.
Chronic theophylline treatment increases adenosine A1, but not A2, receptor binding in the rat brain: An autoradiographic study
Authors: Lupica Carl R. , Berman Robert F. , Jarvis Michael F.
Published: December 2004

Chronic exposure to adenosine receptor antagonists results in an upregulation of brain adenosine A1 receptors as measured by traditional radioligand binding techniques. In the present study, quantitative receptor autoradiography was used to characterize alterations in rat brain adenosine A1 and A2 receptors following the repeated administration of high doses of theophylline. Daily administration of theophylline (75 or 100 mg/kg) markedly increased (125–150% of control) 1 nM [3H]cyclohexyladenosine binding to adenosine A1 receptors in specific cellular layers of the hippocampus, thalamus, and cerebellum with other brain regions showing more moderate increases in binding. By contrast, this chronic theophylline treatment did not produce any significant alterations in the binding of 4 nM [3H]CGS 21680 to adenosine A2 receptors, which were exclusively localized in the striatal region. This apparent differential sensitivity of adenosine receptor subtypes to chronic antagonist treatment suggests a possible intrinsic difference in the regulation of these receptor subtypes which may also be specific to particular brain regions. These results are discussed in relationship to other recent observations, indicating that the pattern of agonist binding to adenosine receptors may be regulated by a differential extent of coupling between adenosine receptors and G‐binding proteins in different brain regions.
Immunohistochemical study of cholecystokinin peptide in rat spinal motoneurons
Authors: Cortés Roser , Åman Katarina , Arvidsson Ulf , Terenius Lars , Frey Peter , Rehfeld Jens F. , Walsh John H. , Hökfelt Tomas
Published: December 2004

With the aid of indirect immunofluorescence histochemistry and sequence specific antibodies a possible localization of cholecystokinin (CCK) peptide in spinal motoneurons has been analyzed. To increase peptide levels, the sciatic nerve was ligated, and the area around the ligation was studied 24 hours later. For comparison, antisera raised against calcitonin gene‐related peptide (CGRP) and substance P were employed.

With CCK specific antisera (directed to the N‐terminal portion of CCK‐8 or the midportion of CCK‐33) accumulation of peptide‐like immunoreactivity (LI) was observed in large, dilated axonal swelling proximal to, but at some distance from, the ligature. Such accumulations were also observed with C‐terminally directed CCK antiserum, but in addition numerous axons of smaller diameter extending up to the ligation contained this type of immunoreactivity. The latter antiserum is thought to cross‐react with CGRP. In fact, this staining pattern was indistinguishable from the one seen after incubation with CGRP antiserum. In contrast substance P‐LI could not be seen in the larger dilated axons but only in large numbers of thinner fibers close to the ligation. Double staining experiments revealed that the large dilations contained both CGRP‐ and CCK‐specific LI. Distal to the ligation CGRP‐ and substance P‐ but no specific CCK‐LI could be observed.

The present findings support the view that CCK mRNA in spinal motoneurons is translated into CCK peptide, at least after axotomy, and that the peptide is transported into the motoneuron axon. However, compared to CGRP the CCK levels are presumably low, and the functional role of CCK peptide in motoneurons remains to be established.
Response to novelty predicts the locomotor and nucleus accumbens dopamine response to cocaine
Authors: Hooks M. Stacy , Jones Graham H. , Smith Amanda D. , Neill Darryl B. , Justice Joseph B.
Published: December 2004

The relationship between a rat's locomotor response to a novel environment and its behavioral and dopaminergic responses to cocaine was examined. Subjects were divided into two groups based on their locomotor response to a novel environment. Subjects who had a novelty response above the median were classified as high responders (HR), while those with a novelty response below the median were classified as low responders (LR). Following administration of cocaine‐HC1 (0, 2.5, 5.0, 10.0, or 15.0 mg/kg), HR rats showed a greater locomotor response than LR rats. Moreover, there was a significant correlation between a subject's locomotor response to the novel environment and the locomotor response to either 10.0 (r = 0.65) or 15.0 (r = 0.92) mg/kg cocaine. In a separate experiment, the extracellular concentration of dopamine in the nucleus accumbens (NACC) was monitored using microdialysis procedures. Following cocaine administration (15.0 mg/kg) HR rats showed a larger NACC dopamine response and greater locomotor activity than LR rats. In addition, there was a threefold greater locomotor activity to dopamine ratio in HR rats than in LR rats. A correlation between a subject's locomotor response to a novel environment and the dopaminergic response to cocaine was also evident. These results suggest that differences in the locomotor response to cocaine can, to some degree, be predicted by a rat's locomotor response to a novel environment, and that variations in dopamine‐dependent mechanisms of the NACC may underlie these individual differences.
Expression of calmodulin‐dependent enzymes in developing rat striatum is not affected by perturbation of dopaminergic systems
Authors: Polli Joseph W. , Kincaid Randall L. , Torris John , Billingsley Melvin L.
Published: December 2004

Transsynaptic regulation is one mechanism that controls expression of several calmodulin (CaM)‐dependent enzymes. This observation and the demonstration that expression of several CaM‐dependent enzymes in developing striatum occurred with a spatial and temporal pattern similar to that seen for dopamine and tyrosine hydroxylase suggested that the nigrostriatal pathway may influence the expression of CaM‐binding proteins (CaM‐BPs) during striatal development. Therefore, the possible role of nigrostriatal dopamine systems regulating the expression of CaM‐dependent enzymes was studied in Sprague‐Dawley rats by using surgical hemitransections of brain, 6‐hydroxydopamine lesions, and chronic haloperidol treatments. Alterations in CaM‐BP expression following perturbation of the developing nigrostriatal tract were analyzed by using immunoblots, biotinylated CaM overlays, and enzyme assays. The extent of nigrostriatal lesions was assessed by using depletion of immunoreactive tyrosine hydroxylase levels in striatum. All there experimental paradigms failed to alter the normal developmental expression of CaM‐dependent enzymes. From these results we conclude that the increased expression of CaM‐dependent enzymes during striatal development is not directly dependent on synaptic input from the nigrostriatal dopamine system.
Effects of neurotensin on midbrain dopamine neurons: Are they mediated by formation of a neurotensin—dopamine complex?
Authors: Shi Wei‐Xing , Bunney Benjamin S.
Published: December 2004

The effects of neurotensin on midbrain dopamine neuron activity were studied in brain slices using single‐unit recording techniques. At low concentrations (0.2–10 nM), neurotensin attenuated dopamine‐induced inhibition without a significant effect on the basal firing rate. At higher concentrations (>10 nM), however, it consistently caused an increase in cell activity. At even higher concentrations (>100 nM), a sudden cessation of cell activity preceded by an increase in firing rate was observed. Whether this effect of neurotensin was due to depolarization inactivation or to a toxic effect of the peptide at high concentrations remains to be determined.

To determine whether the effects of neurotensin were mediated by formation of a neurotensin—dopamine complex, several neurotensin analogues were studied. Neurotensin (8–13), which binds to both neurotensin receptors and dopamine, mimicked the effects of native neurotensin. Neuromedin N, which competes with neurotensin for the same receptor but does not bind to dopamine, also mimicked the effects. However, neurotensin (1–11), which forms a complex with dopamine but is inactive in competing for neurotensin receptors, was ineffective. In addition, the excitatory effect of neurotensin was not attenuated in the presence of dopamine receptor blockade by sulpiride. These results suggest that formation of a neurotensin–dopamine complex may not account for the action of neurotensin on dopamine cells. When combined with the fact that there is a high density of neurotensin receptors on dopamine cells, our results support the suggestion that the observed effects of neurotensin on dopamine neurons are most likely mediated by an activation of neurotensin receptors.
Distribution of cocaine recognition sites in monkey brain: I. In vitro autoradiography with [3H]CFT
Authors: Kaufman Marc J. , Spealman Roger D. , Madras Bertha K.
Published: December 2004

The cocaine analog [3H]CFT ([3H]WIN 35,428) was used to map and characterize cocaine recognition sites in the squirrel monkey brain by quantitative autoradiography. Coronal tissue sections were incubated with 5 nM [3H]CFT to measure total binding or with [3H]CFT in the presence of 30 μM (−)‐cocaine to measure nonspecific binding. High densities of [3H]CFT binding sites were present in dopamine‐rich brain regions, including the caudate nucleus, putamen, nucleus accumbens, and olfactory tubercle. In each of these regions specific binding was greater than 90% of total binding. Several additional brain regions exhibited intermediate densities of [3H]CFT binding, including the substantia nigra, the zona incerta, the amygdala, and the hypothalamus. Low, though measurable levels of binding were observed in the bed nucleus of the stria terminalis, the ventral tegmental area, the medial preoptic area, the pineal, the hippocampus, and thalamic central nuclei. Near‐background levels of binding were found in white matter, cortical regions, globus pallidus, and cerebellum. The pharmacological specificity of [3H]CFT binding in various brain regions was determined in competition studies using [3H]CFT and a range of concentrations of selected monoamine uptake inhibitors. In all brain regions examined, stereoselective inhibition of [3H]CFT binding was observed for the (−) over the (+) isomer of cocaine. For other drugs tested, competition experiments indicated a rank order of potency of GBR 12909 ≥ CFT ≥ bupropion, suggestive of binding of [3H]CFT to elements of the dopamine transport system. The results demonstrate that although densities of [3H]CFT binding sites are highest in the caudate nucleus, putamen, and nucleus accumbens/olfactory tubercle, significant levels of binding can be detected in other brain regions that may contribute to the behavioral and physiological effects of cocaine.
Effects of endogenous dopamine on kinetics of [3H]N‐methylspiperone and [3H]raclopride binding in the rat brain
Authors: Trevor Young L. , Wong Dean F. , Goldman Serge , Minkin Elynne , Chen Catherine , Matsumura Kaname , Scheffel Ursula , Wagner Henry N.
Published: December 2004

Competition by endogenous dopamine with the binding of D2 dopamine receptor ligands may be important in the interpretation of positron emission tomography (PET) neuroreceptor studies. PET studies with N‐methylspiperone (NMSP) have revealed increased D2 dopamine receptors in schizophrenia, whereas studies with raclopride (RAC) have not detected such differences. This may be due, at least in part, to differences in competition with endogenous dopamine for ligand binding. To determine effects of endogenous dopamine on in vivo receptor binding, adult male rats were preinjected with amphetamine and reserpine prior to [3H]NMSP or [3H]RAC. Striatal to cerebellar ratios of ligand binding were determined. To approximate the conditions of a PET study, a kinetic model was employed to examine effects of pharmacologically increasing brain dopamine levels (amphetamine pretreatment) on PET ligand binding. In these experiments, tail veins and arteries were cannulated and kinetic parameters determined from normalized integral plots in rats treated with amphetamine prior to radioligand injection. Both [3H]NMSP (43.5%) and [3H]RAC (41.5%) binding were significantly decreased after amphetamine pretreatment, whereas after reserpine pretreatment [3H]RAC binding was increased (52.7%). Kinetic studies revealed a marked resistance of [3]NMSP to competition with endogenous dopamine released by amphetamine. In contrast, kinetic parameters of [3H]RAC were markedly reduced at all time intervals. This suggests significant differences in competition with endogenous dopamine by [3H]NMSP and [3H]RAC, determined kinetically. These findings may have important implications for the interpretation of PET neuroreceptor studies.
Effects of endogenous dopamine on measures of [18F]N‐methylspiroperidol binding in the basal ganglia: Comparison of simulations and experimental results from PET studies in baboons
Authors: Logan Jean , Dewey Stephen L. , Wolf Alfred P. , Fowler Joanna S. , Brodie Jonathan D. , Angrist Burton , Volkow Nora D. , Gatley S. John
Published: December 2004

The effect of endogenous dopamine on PET measures of radioligand binding is important to the measurement of receptor density (or availability) and neurotransmitter interactions in vivo. We recently reported that pretreatment with amphetamine, a drug which stimulates dopamine release, significantly reduced NMS binding in the baboon brain as determined by the product λk3 dervied from the graphical analysis method for irreversible systems (lambda; is the ratio of the forward to reverse plasma to tissue transport constants and k3 is proportional to receptor density) (Dewey et al.: Synapse 7:324–327, 1991). The purpose of this work is twofold: to evaluate the sensitivity and stability of the analysis method used for the NMS data and from simulation studies which include the competitive effects of dopamine on NMS binding to predict the effect of dopamine on the in vivo PET experiment. Using a measured plasma [18F]‐NMS input function from a control study in a baboon, simulation data was numerically generated explicitly allowing competition between NMS and dopamine in the calculation. This data was analyzed using the same techniques as used for the experimental data and the results were compared to in vitro calculations. The following conclusions were reached: (1) The effect of dopamine on specific binding was found to be greater in vivo than in vitro because the in vitro equilibrium experiment is controlled only by the relative Kd's of tracer and dopamine while the in vivo experiment also depends upon the halftime of tracer in tissue which is controlled by the tissue‐to‐plasma transport constant; (2) Experimental evidence from rodent studies (Seeman et al.: Synapse 3:96–97, 1989) and the agreement between PET studies (Wong et al.: Science 225:728–731, 1984) in schizophrenics suggest that NMS is not likely to be affected by normal levels of endogenous dopamine. From the calculations reported here the effective in vivo Kd of dopamine for the NMS binding site would have to be on the order of or greater than 100 nM, assuming a synaptic dopamine concentration of 20 nM, in order that this concentration of dopamine have little effect on NMS binding.
Excitatory responses to serotonin (5‐HT) in neurons of the rat piriform cortex: Evidence for mediation by 5‐HT1C receptors in pyramidal cells and 5‐HT2 receptors in interneurons
Authors: Sheldon Priscilla W. , Aghajanian George K.
Published: December 2004

As a prerequisite to pharmacological analysis of the excitatory effects of serotoinin (5‐HT) on piriform pyramidal cells and interneurons, this study first examined the physiological characteristics of these two cell types. Intracellular recordings confirmed that the subpopulation of 5‐HT‐activated cells located at the border of layers II and III and indeed interneurons. Voltage clamp recordings in pyramidal cells showed that the increase in excitability produced by 5‐HT in these cells was the result of voltage‐ and Ca2+‐dependent outward currents with the characteristics of IM and IAHP.

Pharmacological studies were designed to discriminate 5‐HT2 from 5‐HT1C responses in interneurons and pyramidal cells of piriform cortex. The 5‐HT antagonist spiperone, which has a much higher affinity for 5‐HT2 receptors than for 5‐HT1C receptors, blocked the excitatory effect of 5‐HT at lower concentrations in interneurons (IC50 = 31 nM) than in pyramidal cells (IC50 = 2.1 μM). Similarly, ritanserin, a drug which also has a higher affinity for 5‐HT2 than 5‐HT1C receptors, blocked the effect of 5‐HT at lower concentrations in interneurons (IC50 = 400 nM) than in pyramidal cells (IC50 = 8.1 μM). In contrast, LY 53857, an antagonist with higher affinity for 5‐HT1C than for 5‐HT2 receptors, blocked the effect of 5‐HT at lower concentrations in pyramidal cells (IC50 = 26 nM) than in interneurons (IC50 = 364 nM). The 5‐HT1C partial agonist/5‐HT2 antagonist mCPP produced agonist‐like effects in only 66% of pyramidal cells tested indicating that not all pyramidal cells may express 5‐HT1C receptors.

In that both spiperone and ritanserin have higher affinity for 5‐HT2 receptors than for 5‐HT1C receptors and LY 53857 has a higher affinity for 5‐HT1C receptors than for 5‐HT2 receptors, these data suggest that in piriform cortex excitatory effects of 5‐HT are mediated by 5‐HT1C receptors in pyramidal cells and by 5‐HT2 receptors in interneurons.