Administration Of DSP-4 Research Articles

Localization of a glutathione-dependent dehydroascorbate reductase within the central nervous system of the rat

In this study, we describe for the first time the occurrence, within the central nervous system of the rat, of a dehydroascorbate reductase analogous to the one we recently described in the liver. Dehydroascorbate reductase plays a pivotal role in regenerating ascorbic acid from its oxidation product, dehydroascorbate. In a first set of experiments, we showed that a dehydroascorbate reductase activity is present in brain cytosol; immunoblotting analysis confirmed the presence of an immunoreactive cytosolic protein in selected brain areas. Immunotitration showed that approximately 65% of dehydroascorbate reductase activity of brain cytosol which was recovered in the ammonium sulphate fraction can be attributed to this enzyme. Using immunohistochemistry, we found that a variety of brain areas expresses the enzyme. Immunoreactivity was confined to the gray matter. Amongst the several brain regions, the cerebellum appears to be the most densely stained. The enzyme was also abundant in the hippocampus and the olfactory cortex. The lesion of norepinephrine terminals following systemic administration of DSP-4 markedly decreased immunoreactivity in the cerebellum. Apart from the possible co-localization of the enzyme with norepinephrine, the relative content of dehydroascorbate reductase in different brain regions might be crucial in conditioning regional sensitivity to free radical-induced brain damage. Given the scarcity of protective mechanisms demonstrated in the brain, the discovery of a new enzyme with antioxidant properties might represent a starting-point to increase our knowledge about the antioxidant mechanisms operating in several central nervous system disorders.

Astroglial and vascular interactions of noradrenaline terminals in the rat cerebral cortex.

Noradrenaline (NA) has been shown to influence astrocytic and vascular functions related to brain homeostasis, metabolism, local blood flow, and blood-brain barrier permeability. In the current study, we investigate the possible associations that exist between NA-immunoreactive nerve terminals and astrocytes and intraparenchymal blood vessels in the rat frontoparietal cortex, both at the light and electron microscopic levels. As a second step, we sought to determine whether the NA innervation around intracortical microvessels arises from peripheral or central structures by means of injections of N-(2-chloroethyl-N-ethyl-2-bromobenzylamine) (DSP-4), a neurotoxin that specifically destroys NA neurons from the locus ceruleus. At the light microscopic level, 6.8% of all NA-immunoreactive nerve terminals in the frontoparietal cortex were associated with vascular walls, and this perivascular noradrenergic input, together with that of the cerebral cortex, almost completely disappeared after DSP-4 administration. When analyzed at the ultrastructural level in control rats, NA terminals in the neuropil had a mean surface area of 0.53 +/- 0.03 micron2 and were rarely junctional (synaptic incidence close to 7%). Perivascular terminals (located within a 3-micron perimeter from the vessel basal lamina) counted at the electron microscopic level represented 8.8% of the total NA terminals in the cortical tissue. They were smaller (0.29 +/- 0.01 micron2, P < 0.05) than their neuronal counterparts and were located, on average, 1.34 +/- 0.08 microns away from intracortical blood vessels, which consisted mostly of capillaries (65%). None of the perivascular NA terminals engaged in junctional contacts with surrounding neuronal or vascular elements. The primary targets of both neuronal and perivascular NA nerve terminals consisted of dendrites, nerve terminals, astrocytes, and axons, whereas in the immediate vicinity (0.25 micron or less) of the microvessels, astrocytic processes represented the major target. The results of the current study show that penetrating arteries and intracortical microvessels receive a central NA input, albeit parasynaptic in its interaction, originating from the locus ceruleus. Particularly, they point to frequent appositions between both neuronal and perivascular NA terminals and astroglial cells and their processes. Such NA neuronal-glial and neuronal-glial-vascular associations could be of significance in the regulation of local metabolic and vascular functions under normal and pathologic situations.

Overflow of noradrenaline and dopamine in frontal cortex after [N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine] (DSP-4) treatment: in vivo microdialysis study in anaesthetized rats.

The changes in the extracellular concentration of endogenous noradrenaline and dopamine in the frontal cortex following pretreatment with noradrenergic neurotoxin DSP-4 [N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine] were studied by in vivo microdialysis in rats anaesthetized with chloral hydrate. Noradrenaline and dopamine levels in frontal cortex were detected only when the uptake inhibitor, nomifensine (10 microM) was present in dialysis fluid. Under those conditions, the Na+ channel agonist veratridine and a depolarising concentration of potassium chloride (60 mM), applied locally through the microdialysis probe, increased the overflow of noradrenaline. Tetrodotoxin had an opposite effect. These results indicate that most of the noradrenaline probably arose from exocytotic release. Noradrenaline efflux in the frontal cortex of DSP-4 pretreated rats (52 +/- 6.1 fmol/sample) did not differ significantly from that of the control animals (69 +/- 4.9 fmol/sample). Dopamine efflux was not changed either (64 +/- 9.6 and 62 +/- 3.9 fmol/sample, respectively). The alpha 2-adrenoceptor antagonist, atipamezole (3 mg/kg i.p.), increased the overflow of noradrenaline in the frontal cortex of saline-treated rats by 100%, whereas in DSP-4 treated rats the increase was only around 30%. The overflow of dopamine was not changed under the conditions described. The effect of atipamezole in DSP-4 treated rats may be of smaller magnitude due to the diminished pool of releasable noradrenaline or due to a downregulation of presynaptic alpha 2-adrenoceptors in the frontal cortex. The perfusion of 60 mM KCl at the end of the experiment unexpectedly produced equivalent increases in noradrenaline and dopamine content in dialysates of both vehicle and DSP-4 treated rats. We conclude that the uptake inhibitor, nomifensine, and atipamezole, which had a stronger effect in vehicle-treated animals, reduced the effect of KCl-stimulation and masked the true difference in changes of noradrenaline efflux. Post-mortem tissue concentrations of noradrenaline 7 days after DSP-4 administration (50 mg/kg) were significantly reduced in the frontal cortex (54%), hippocampus (62.5%) and to lesser extent in the hypothalamus (27%) as compared to vehicle-treated rats. Dopamine and 3,4-dihydroxyphenylacetic acid concentrations were not changed confirming the efficacy and selectivity of the DSP-4 lesion. These results demonstrate that one week after DSP-4 treatment the extracellular levels of noradrenaline and dopamine as assessed by in vivo microdialysis are not changed in the frontal cortex, but atipamezole-stimulated release of noradrenaline is decreased in DSP-4 treated rats.

Region- and Neurotransmitter-dependent Species and Strain Differences in DSP–4–induced Monoamine Depletion in Rodents

The neurotoxin N-(-2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) is commonly used as a chemical tool to induce selective denervation of noradrenergic terminals arising from the locus coeruleus and to study the molecular mechanisms underlying degeneration of central noradrenergic axons in rodents. Monoamine depletion in different rodent species after DSP-4 is generally assumed to occur with a similar pattern. To verify this assumption, in the present study we evaluated the different patterns of monoamine depletion produced by DSP-4 in different brain regions of two different strains of mice and rats 3, 7 and 14 days after DSP-4 administration. In this report, we show that there are evident species and strain differences concerning the pattern of norepinephrine depletion in various brain regions. Moreover, serotonin levels are fully preserved following DSP-4 in mice, whereas there is a significant serotonin decrease in specific brain regions after the same dose of DSP-4 in rats. Apart from disclosing species and strain variability among rodents in neurotoxin-induced monoamine depletion, these findings suggest that DSP-4 should be considered as a different neurotoxin, depending on the species and strain in which it is administered.

Neurotoxic effects of DSP-4 on the noradrenergic system of the goldfish brain.

The substance N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) is a neurotoxin with selective and long-lasting effects on the noradrenergic (NA) neurons of mammalian brains. The present study examines the effects of this toxin on the noradrenergic system of the goldfish brain. Single doses (50 mg/kg body weight) of DSP-4 reduce the immunoreactivity of the NA synthesizing enzyme dopamine-beta-hydroxylase (DBH), as revealed by immunohistochemistry 7 and 12 days after toxin administration. The depletion involves the DBH-positive fibres and spares the DBH-positive cell bodies. Dopamine-beta-hydroxylase immunoreactivity, 40 days after toxin administration, showed a complete recovery. Ultrastructural investigations confirmed that DSP-4 toxicity affects only nervous fibres and terminals, sparing cell bodies. Administration of DSP-4 also produced a marked decrease of noradrenaline (NA) levels in the goldfish brain, seven days later, while dopamine (DA) and serotonin (5-HT) levels were unaffected by toxin injection. The reduction of NA levels induced by DSP-4 was prevented by the concomitant administration of the NA uptake inhibitor desipramine. Noradrenaline levels measured 40 days after toxin administration show that DSP-4 toxicity was completely reversed. The results suggest a pronounced plasticity of the noradrenergic system in the goldfish brain.

Central DSP-4 treatment decreases norepinephrine levels and courtship behavior in male zebra finches

In zebra finches, gonadal steroids activate male courtship, including singing, and also strongly modulate norepinephrine (NE) levels and turnover in brain areas regulating courtship behavior. In a previous study, systemic administration of DSP-4 caused significant decreases in courtship singing. These behavioral decrements were correlated with the degree of NE depletion in several vocal control nuclei. In the present study, we attempted to further decrease brain NE levels while minimizing systemic effects by infusing DSP-4 directly into the third ventricle. DSP-4 treatment significantly reduced NE levels in three of six vocal control nuclei and both hypothalamic nuclei sampled without significantly altering dopamine or serotonin levels in any areas. DSP-4-treated males took longer to begin singing and performed fewer song bouts and courtship displays. Interestingly, behavioral deficits were limited to courtship song displays, other behavior patterns, including female-directed behaviors like approach and follow, were unaffected by DSP-4 treatment. DSP-4 treatment appeared to affect singing behavior by causing deficits in initial attentiveness to females and initiation of singing rather than by affecting song structure.

Impaired exploratory behaviour after DSP-4 treatment in rats: implications for the increased anxiety after noradrenergic denervation.

The effect of DSP-4, a neurotoxin selectively affecting the projections of the locus coeruleus, upon exploratory behaviour of rats was studied in a complex exploration test, including a choice between open and enclosed areas and both inquisitive and inspective exploration elements. One week after DSP-4 (50 mg/kg i.p.) administration, the neurotoxin-treated rats did not explore a novel open area to any extent on the first exposure to the apparatus; however, on the third day of testing, these animals began to explore the area and the novel objects. Diazepam (0.5 mg/kg) treatment did not change the behaviour of control rats, but significantly increased the exploratory activity of the DSP-4-treated animals. LY 288513, a selective CCKB receptor antagonist (0.01 mg/kg), prevented the increase in exploratory activity in the DSP-4-treated rats, but increased the exploratory activity of controls on the third exposure to the test situation. The results of this investigation suggest that DSP-4 treatment reduces neotic behaviour by increasing neophobia rather than by decreasing exploratory drive. The divergence reported between the relative potencies of CCKB receptor antagonists in exploratory activity and other anxiety tests may in part be due to the effects of these drugs on exploratory drive.

Impaired exploratory behaviour after DSP-4 treatment in rats: implications for the increased anxiety after noradrenergic denervation

The effect of DSP-4, a neurotoxin selectively affecting the projections of the locus coeruleus, upon exploratory behaviour of rats was studied in a complex exploration test, including a choice between open and enclosed areas and both inquisitive and inspective exploration elements. One week after DSP-4 (50 mg/kg i.p.) administration, the neurotoxin-treated rats did not explore a novel open area to any extent on the first exposure to the apparatus; however, on the third day of testing, these animals began to explore the area and the novel objects. Diazepam (0.5 mg/kg) treatment did not change the behaviour of control rats, but significantly increased the exploratory activity of the DSP-4-treated animals. LY 288513, a selective CCK B receptor antagonist (0.01 mg/kg), prevented the increase in exploratory activity in the DSP-4-treated rats, but increased the exploratory activity of controls on the third exposure to the test situation. The results of this investigation suggest that DSP-4 treatment reduces neotic behaviour by increasing neophobia rather than by decreasing exploratory drive. The divergence reported between the relative potencies of CCK B receptor antagonists in exploratory activity and other anxiety tests may in part be due to the effects of these drugs on exploratory drive.

Recovery of central noradrenergic neurons one year after the administration of the neurotoxin DSP4

The long-term effects of the systemic administration of DSP4 (N-(2-chloroethyl)N-ethyl-2-bromobenzylamine hydrochloride), a selective noradrenergic neurotoxin, on the endogenous levels of monoamines and their metabolites and on alpha- and beta-adrenoceptors in selected brain regions of the rat were examined. After 7 days, DSP4 caused a marked reduction (about 80%) of endogenous noradrenaline levels in locus coeruleus-innervated regions. At 90, 240 and 300 days after DSP4 injection, a partial and gradual recovery (50%, 41% and 25% of control values, respectively) of the noradrenaline cortical levels was evident. One year after DSP4 administration, brain regional noradrenaline stores were almost completely recovered. No changes in 5-hydroxytryptamine levels were observed in the three time intervals, but a mild decrease in cortical and hippocampal 5-hydroxyindolacetic acid levels was found 7 days after DSP4 injection. Following the profound noradrenaline depletion seen at 7 days, the cerebral cortical density of alpha 1-, alpha 2- and beta-adrenoceptors was significantly increased. Assessment of adrenergic receptors in cerebral cortex at 365 days after DSP4 injection, indicated that alpha 1- and alpha 2-adrenoceptor densities did not differ from control values; however, the density of beta-adrenoceptors remained increased. No changes were observed in the affinities of the three types of adrenoceptors studied. These results indicate that after a selective noradrenergic denervation induced by DSP4, there is a slow and gradual recovery of noradrenaline stores and of alpha 1- and alpha 2-adrenoceptor populations, suggesting a possible regrowth and/or collateral sprouting of noradrenergic terminals.

Disruptive influence of norepinephrine depletion on sensory preconditioning, but not first-order conditioning, in preweanling rats

Learning and memory processes are known to be influenced by the action of norepinephrine (NE). The present study evaluated the influence of neonatal administration of the NE neurotoxin DSP4 on sensory preconditioning (SPC) in 16- and 28-day-old rats. Rats were subcutaneously administered 50 mg/kg DSP4 or saline within 24 h after birth and tested at 16 or 28 days in two experiments. The results from Experiment 1 indicated that nondepleted 16-day-old rats exhibited strong SPC, whereas this conditioning was disrupted by neonatal administration of DSP4. No SPC was seen in either neonatal treatment group at 28 days of age, confirming previous reports of an ontogenetic decline in SPC. In Experiment 2, no effects of DSP4 were observed on first-order conditioning at either age. The brains of representative subjects were separated into forebrain, cerebellum, and brain stem samples and were analyzed to determine the extent of NE depletion. This analysis showed that DSP4 produced a marked reduction in NE concentration in the forebrain, but not in the cerebellum or brain stem. The results of this study suggest a role for forebrain NE in SPC in preweanling rats. This modulatory noradrenergic influence appears to be exerted on the formation of associations between two relatively neutral stimuli during the preexposure phase, given the absence of an effect of DSP4 on primary conditioning.

Conflict behavior in maudsley reactive and nonreactive rats: Effects of noradrenergic neuronal destruction

The present studies were designed to examine the effects of treatment with the noradrenergic neurotoxin N-(2-chloroethyl)- n-ethyl-2-bromobenzylamine HCl (DSP4; 65 mg/kg, IP) on conflict behavior in the Maudsley reactive (MR) and nonreactive (MNRA) rat strains. In daily 10-min sessions, water-restricted rats were trained to drink water from a tube that was ocassionally electrified; electrification was signaled by the presence of a tone (7-s duration; ISI = 30 s). Consistent with previous reports, the number of shocks accepted by rats of the MR and MNRA strains did not differ initially, but MNRA rats exhibited a dramatic increase in punished responding relative to their MR counterparts over the course of several weeks of conflict testing. This MR vs. MNRA strain difference in punished responding did not exhibit extinction following discontinuation of CSD conflict behavior testing for a period of 6 weeks. Whether it was administered after conflict training or before, DSP4 treatment did not reduce the MR vs. MNRA strain difference in conflict behavior; rather, DSP4 treatment tended to increase the magnitude of the MR vs. MNRA difference in conflict behavior. The effects of DSP4 on norepinephrine (NE) and 5-hydroxytrypamine (5-HT) concentrations in the pons medulla region were determined in one group of conflict-experienced MR and MNRA rats (35 weeks after administration) and in a second group of naive MR and MNRA rats (3 weeks after administration). There were no MR vs. MNRA strain differences in NE or 5-HT concentrations in vehicle-treated rats. DSP4 treatment significantly reduced NE, but not 5-HT, concentrations when compared to control values; rats that were sacrificed 3 weeks following DSP4 administration exhibited a greater NE depletion than did rats sacrificed 35 weeks after DSP4 administration. Finally, there were no significant correlations between pons medulla region NE concentrations and conflict behavior in either strain alone or when the data from the two strains were combined. The present results are not consistent with the hypothesis that the MR vs. MNRA strain difference in conflict behavior is the result of strain differences in brain NE function.

Neurotoxins and monoamine oxidase inhibition: new aspects.

Recent clinical studies suggest that selegiline (L-deprenyl) is useful in retarding the progress of Parkinson's disease, an effect that may be related to its inhibition of monoamine oxidase type B (MAO-B). Selegiline is also reported to prevent the toxic effects of the noradrenergic neurotoxin, N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4). This article reviews recent studies on the role of MAO-B and its inhibition in this neuroprotective action of selegiline. Male C57Bl/6 mice were given DSP-4 (50 mg/kg) 1 h, 24 h, or 4 days after the administration of selegiline (10 mg/kg) or the selective MAO-B inhibitor MDL 72974 (1.25 mg/kg) and then killed 1 week later for the assay of norepinephrine in the hippocampus. The MAO-B-inhibiting effects of selegiline or MDL 72974 were also determined after these same intervals. Selegiline and MDL 72974 produced comparable degrees of enzyme inhibition 1 h (> 95%), 24 h (> 90%), or 4 days (> 70%) after their administration. Given 1 h before, selegiline totally blocked the norepinephrine-depleting effects of DSP-4, but this protection declined sharply when 24 h or 4 days was allowed to elapse between selegiline and DSP-4 administration. MDL 72974 failed to protect at any time point. In vitro, no activity was observed when DSP-4 was used as a substrate for MAO. All of these findings suggest that the ability of selegiline to protect against DSP-4-induced neuronal degeneration does not depend on its inhibition of MAO-B.

Reversal of learning and memory impairments following lesion of the nucleus basalis magnocellularis (NBM) by concurrent noradrenergic depletion using DSP4 in the rat

In the following study the behavioural effects of simultaneous lesion of the nucleus basalis magnocellularis (NBM) using ibotenic acid and noradrenergic depletion following a single i.p. administration of DSP4 (50 mg/kg) were examined in the rat. NBM lesion induced a deficit in acquisition of a reinforced T-maze alternation task, a working memory adaptation of a spatial navigation task in a water maze and 24 h retention in a passive avoidance task compared to sham controls. No effect of the lesion on a reference memory version of spatial navigation in a water maze task was found. Animals that received a combination of NBM lesion and DSP4 treatment showed no impairment on any of the tasks that were impaired by NBM lesion alone. This indicates a reversal of the learning and memory deficits consequent to NBM lesion by simultaneous noradrenergic depletion. NBM lesion induced a significant reduction in cholineacetyltransferase activity in the frontal cortex, and DSP4 induced a significant decrease in noradrenaline concentration in occipital cortex and hippocampus, confirming the effects of these treatments. These results suggest an interaction between central noradrenergic and cholinergic systems in learning and memory processes.

Effects of chronic catecholamine depletions on muscarinic M1-receptor and its mRNA in rat brain

In order to compare the effects of total catecholamine (CA) or noradrenaline (NA) depletions on cholinergic systems, and the mechanisms of receptor regulation in various brain regions, the regional changes in the levels of acetylcholine (ACh), M1-receptor (M1-R) binding, and M1-R messenger RNA (mRNA) were mainly examined in rats which had received either repeated reserpine treatment or a single injection of the selective noradrenergic neurotoxin N-2-chloroethyl- N-ethyl-2-bromobenzylamine (DSP-4). The levels of dopamine (DA), its metabolites, NA, binding to both D1 and D2 sites, and the mRNA encoding the D2 receptor were also measured. Administration of reserpine (0.5 mg/kg/day, s.c.) for 2, 7 and 14 days depleted DA and NA in virtually all brain regions, while the short-term treatment increased DA metabolites in the striatum (at 2 days) and basal forebrain (at both 2 and 7 days). Administration of DSP-4 (50 mg/kg, i.p.) resulted in a specific loss of NA in the brain 10 days after the injection. These DSP-4 treated rats showed no change in the levels of ACh or M1-R except for an increase in ACh in the frontal cortex. In contrast, numerous changes in cholinergic indices were seen in the reserpine treated groups, and these changes varied from region to region of brain and with the length of drug treatment. In the striatum, ACh levels were increased in rats treated for 2 or 7 days but were normal after 14 days. M1-Rs were decreased at 14 days. These changes suggest that striatal DA, initially released by reserpine, inhibits the release of ACh from striatal cholinergic interneurons, while prolonged depletion of DA relieves this inhibition, leading to a subsequent down-regulation of M1-Rs. In the frontal cortex, ACh and M1-R levels were all decreased by reserpine treatment for 2 or 7 days, 7and the M1-Rs remained depressed at 14 days. In the basal forebrain, which contains the cholinergic cells that project to the cortex, DA metabolism was increased by 2 or 7 day reserpine treatment. This increased DAergic activity in the basal forebrain may facilitate cholinergic neurons, causing increased release of ACh in the frontal cortex. This, in turn, may lead to a down-regulation of the M1-Rs in that region. The levels of mRNAs encoding M1-Rs were increased in the striatum and frontal cortex by reserpine treatment, despite the decreases in the M1-Rs themselves. This indicates that the down-regulation of M1-Rs in both regions may be due to an increase in receptor degradation rather than a decrease in receptor production. This contrasts with the mechanism of up-regulation of the striatal D2 receptor in reserpinized rats which correlates with increased expression of its mRNA. The present results indicate that cholinergic neurons in the striatum and basal forebrain are both regulated by DAergic neurons, but in different fashions.

Lesions of noradrenergic neurons in rats with spontaneous generalized non-convulsive epilepsy

The role of noradrenergic neurons in the control of a spontaneous generalized non-convulsive epilepsy (GNCE) was investigated. In rats with genetic spontaneous absence seizures, we produced lesions using 2 neurotoxins: 6-hydroxydopamine (6-OHDA) and N-(2-chloroethyl)- N-ethyl-2-bromobenzylamine (DSP4). Lesions of noradrenergic neurons were made either in pups by neonatal 6-OHDA intraperitoneal (i.p.) injection (2 × 100 mg/ kg) or in adult rats by i.p. administration of DSP4 (60 mg/kg) or bilateral microinjection of 6-OHDA in the locus coeruleus (LC) (4μg/μl, 2μl/side). Effectiveness of the lesions was controlled by measuring dopamine (DA) and noradrenaline (NA) contents in the brains. Neonatal 6-OHDA administration did not lead to any difference in seizures in adult animals, compared with control rats. DSP4 injections and LC lesions with local injections of 6-OHDA produced a transient increase of the seizures. Within one to two weeks, the seizure duration went back to prelesion levels. No seizure occurred when the same lesions were performed in non epileptic rats. These results suggest that NA is not involved in the genesis of this generalized non-convulsive epilepsy; they confirm that NA participates in the control of seizures in this model, but the rapid development of compensatory mechanisms shows that this control is not critical.

Noradrenergic Control of Thalamic Oscillation: the Role of alpha-2 Receptors.

The effects of alpha-adrenergic drugs on neocortical high voltage spike and wave spindles (HVS), reflecting thalamic oscillation, was investigated in freely moving rats. HVS occurred spontaneously in the awake but immobile animal. Peripheral administration of the alpha-1 antagonist, prazosin and alpha-2 agonists, xylazine and clonidine increased the incidence and duration of HVS in a dose-dependent manner. The alpha-2 antagonist, yohimbine and the tricyclic antidepressants, desipramine and amitriptyline, significantly decreased the incidence of the neocortical HVS. Bilateral microinjections of the alpha-2 agonists into the nucleus ventralis lateralis area of the thalamus, but not into the hippocampus or corpus callosum, was as effective as peripheral injection of these drugs. Xylazine was most effective in Fischer 344 rats that display high spontaneous rate of HVS and less effective in the Sprague - Dawley and Buffalo strains. The HVS-promoting effect of clonidine was antagonized by prior intrathalamic injection of the alpha-2 antagonist, yohimbine. The amplitude of the HVS was increased by picomole amounts of unilaterally-injected clonidine. Neurotoxic destruction of the thalamopetal noradrenergic afferents by intracisternal or intrathalamic injection of 6-hydroxydopamine, but not by peripheral administration of DSP-4, increased the incidence of HVS. Importantly, intrathalamic administration of xylazine continued to induce HVS after destroying the thalamic noradrenergic terminals. Following downregulation of the alpha-2 adrenoceptors by chronic administration (3 weeks) of amitriptylene the incidence of HVS decreased and the effectiveness of intrathalamic xylazine on the induction of HVS was significantly reduced. Based on these findings, we suggest that a major action of alpha-2 adrenergic drugs on thalamic oscillation may be mediated by postsynaptic alpha-2 adrenoceptors located on the thalamocortical neurons. We hypothesize that noradrenaline in the thalamus has a dual effect on the relay cells: blocking and promoting thalamic oscillation via alpha-1 and alpha-2 receptors, respectively. The final physiological effect is assumed to be a function of the relative density and affinity of these adrenergic receptor subtypes.

Protection against DSP-4-induced neurotoxicity by deprenyl is not related to its inhibition of MAO B

Clinical studies suggest that deprenyl may retard the progression of Parkinson's disease, an effect that may be related to its monoamine oxidase (MAO) inhibiting properties. Deprenyl also protects against the neurodegenerative effects of the noradrenergic toxin DSP-4. In this study we investigated the role of MAO B inhibition in this protection. C57BL/6 mice were given DSP-4 (50 mg/kg i.p.) 1 h, 24 h or 4 days after the administration of deprenyl (10 mg/kg i.p.) or the selective MAO B inhibitor MDL 72974 (1.25 mg/kg), and then killed 1 week later for assay of hippocampal norepinephrine. The MAO B inhibiting effects of deprenyl or MDL 72974 were also determined after these same intervals of time. Deprenyl and MDL 72974 produced comparable degrees of enzyme inhibition 1 h (> 95%), 24 h (> 90%) or 4 days (> 70%) after their administration. Given 1 h before, deprenyl totally blocked the norepinephrine-depleting effects of DSP-4, but this protection declined sharply when 24 h or 4 days was allowed to elapse between deprenyl and DSP-4 administration. MDL 72974 failed to protect at any time point. In vitro, we detected no activity using DSP-4 as a substrate for MAO. These findings suggest that the ability of deprenyl to protect against DSP-4-induced neuronal degeneration may not depend on its MAO B inhibiting properties.

Complex maze performance in rats: Effects of noradrenergic depletion and cholinergic blockade.

Noradrenergic (NA) involvement in acquisition (AQ) and retention (RET) of a shock-motivated 14-unit T-maze after intraperitoneal (ip) administration of DSP4, a neurotoxin that depletes forebrain NA, was evaluated in 2 experiments using 3-month-old male F-344 rats. Depletion of cortical NA level in DSP4-treated rats was verified by neurochemical assay and by latency to postdecapitation clonus reflex. In Experiment 1, DSP4 (50 mg/kg ip) rats did not differ from saline-treated (SAL) rats on any measure of maze performance (errors, alternation errors, run time, shock duration, or shock frequency) during AQ (DSP4 2 weeks prior to testing) nor during a RET test 2-3 weeks later (DSP4 immediately after the last AQ trial or 1 week after AQ). In Experiment 2, DSP4 and scopolamine (SCOP), a cholinergic (ACh) antagonist, were administered in combination to test for an NA-ACh interaction. DSP4 (50 mg/kg ip 2 weeks prior to AQ) again had no effect on AQ performance nor did DSP4 interact with either dose of SCOP (0.25 or 0.5 mg/kg ip 30 min prior to AQ) to further impair performance. Similar to previous studies SCOP impaired all measures of maze performance except shock duration. These experiments provide further evidence for involvement of ACh systems in the learning of this task and in the age-related impairments previously observed, but they suggest that central NA systems may not be involved directly or interactively with ACh systems required for efficient performance in this maze.

Acute action of DSP-4 on central norepinephrine axons: biochemical and immunohistochemical evidence for differential effects.

Previous immunohistochemical studies of the long-term effects of the noradrenergic neurotoxin DSP-4 have demonstrated a remarkably selective vulnerability of norepinephrine (NE) axons of the locus coeruleus (LC). NE axons originating in non-LC NE neurons appear to be largely resistant to the neurotoxic action of DSP-4. We conducted this study to evaluate the acute effects of DSP-4 on NE axons in four different brain regions: cerebral cortex, cerebellum, ventral forebrain, and hypothalamus. NE levels were determined by high-performance liquid chromatography (HPLC) 6 and 24 hr and 14 days after DSP-4 administration. NE axons in these brain regions were visualized in brain sections at 6 and 24 hr after drug treatment, using a specific antiserum to NE. HPLC assays revealed profound reductions of NE levels in cerebral cortex and cerebellum, but only minor decreases in ventral forebrain and hypothalamus. NE immunohistochemistry showed dramatic differences in the acute effects of DSP-4 on NE axon staining: nearly complete loss of staining in cortex and cerebellum, in contrast to an almost unchanged staining pattern in ventral forebrain and hypothalamus. This study demonstrates that NE immunohistochemistry is a valuable tool to assess the acute effects of DSP-4 on NE axons in different brain regions. The results provide the first direct evidence that NE axons are not uniformly acted on by DSP-4 and suggest that the acute effects of DSP-4 are restricted to LC axons.

P-178 - Effects of prenatel administration of DSP4 on growth, locomotor activity, and brain monoamine contents in offspring mice

P-178 - Effects of prenatel administration of DSP4 on growth, locomotor activity, and brain monoamine contents in offspring mice