6-hydroxydopamine Treatment Research Articles

Effects of 6-hydroxydopamine treatment on locomotor response to l-DOPA in neonatal rats

Error scores on the Farnsworth-Munsell 100-hue test were partitioned into those representing red-green and those representing blue-yellow losses. Data from two groups of normal observers were used. One group showed results characteristic of published norms; one group showed superior performance. Both observers showed a correlation between red-green and blue-yellow scores indicative of a strong performance factor in this test. The difference between blue-yellow and red-green scores eliminates their correlated variance and allows evaluation of the axis. Both groups showed an increase in difference scores, with age indicating development of a blue-yellow axis. This increase was significant for the observers characteristic of the norms. We suggest cutoff scores to allow a decision as to whether a given patient shows a blue-yellow or red-green axis.

Presence of neuropeptide Y in the rat seminal vesicle and its effects on noradrenaline‐ and nerve‐induced contractions

1. Immunohistochemical and functional studies have been performed to localize and determine the effects of neuropeptide Y (NPY) in the rat seminal vesicle. 2. An abundant presence of NPY-immunoreactive nerves, mainly concentrated in the smooth muscle layer of the seminal vesicle was found. Chronic 6-hydroxydopamine treatment (four doses of 50 mg kg-1 i.p. on days 1, 2, 4 and 6; rats killed one week after the last injection) led to a large reduction but not abolition of the NPY-immunoreactivity. 3. NPY (1-250 nM) did not affect the resting tone of the seminal vesicle. 4. The seminal vesicle was contracted by electrical field stimulation (EFS) and by exposure to 5 microM noradrenaline (NA). These contractions were abolished by phentolamine (1 microM). Tetrodotoxin (0.5 microM) abolished EFS-evoked contractions but did not affect NA-evoked contractions. 5. Seminal vesicles, from animals chronically-treated with reserpine (5 mg kg-1 i.p. on days 1 and 2; rats killed on day 3), were contracted by NA but not by EFS. 6. NPY (0.25-250 nM), concentration-dependently, inhibited EFS-evoked contractions by up to 70% maximum inhibition. Contractions evoked by EFS with short trains of pulses were inhibited by NPY to a greater degree than those with longer trains. 7. NPY had no significant effect on NA-evoked contractions. 8. These data provide strong evidence that the motor transmission in rat seminal vesicle is predominantly if not exclusively, adrenergic. It is further concluded that a rich NPY-containing innervation is present in the smooth muscle layer of rat seminal vesicle. The primary effect of NPY is modulation of adrenergic motor transmission by a prejunctional inhibition of NA release.

Lack of effect of bilateral locus coeruleus lesion and antidepressant treatment on gamma-aminobutyric acidB receptors in the rat frontal cortex

The aim of the present study was to investigate whether a disturbance of the central noradrenergic (NA) system could cause changes in gamma-aminobutyric acidB (GABAB) receptors in the rat frontal cortex. Manipulation of the NA projection to the frontal cortex was achieved by bilateral lesion of the locus coeruleus with 6-hydroxydopamine (6-OHDA) or chronic treatment with the NA reuptake blocker and antidepressant drug, desipramine. Precautions were taken to ensure that the GABAB receptor assay was performed optimally and was not confounded by the presence of endogenously generated GABA. The results show conclusively that manipulation of the NA projection did not result in any significant change in the number (Bmax) or affinity (Kd) of GABAB receptors in the frontal cortex. These results do not support the hypothesis that hypoactivity of the central NA system can lead to changes in cortical GABAB receptors and that antidepressant drugs act by increasing GABAB receptor binding in the frontal cortex.

Adrenal medullary adaptations and cardiovascular regulation after 6-hydroxydopamine treatment in rats

Experiments were conducted to assess the pattern of activation of adrenal medullary adrenaline (ADR) and noradrenaline (NA) chromaffin cells following damage to sympathetic neurons with 6-hydroxydopamine (6-HDA, 100 mg/kg per day for 7 days, s.c.). 6-HDA treatment resulted in a sustained elevation in urinary ADR excretion, suggesting that the adrenal medulla was continuously stimulated. Further evidence of neural stimulation of the adrenal medulla was demonstrated by a persistent elevation of adrenal medullary dopamine (DA) content, an indicator of catecholamine synthesis. The increase in synthetic rate was associated with a significantly augmented medullary NA and ADR content. However, NA content was elevated after only 1 day of treatment, whereas, ADR became elevated after the second day, a finding that suggests a differential activation of the two populations of chromaffin cells. Cardiac and kidney NA contents were reduced by 97 and 87%, respectively, and BP increments to stimulation of the thoracolumbar spinal cord of pithed rats (0.5 to 16 Hz) were attenuated by more than 75% by 6-HDA. Moreover, the residual BP responses were abolished by adrenalectomy. Thus the results indicate that the changes in adrenal CA disposition may be related to a greater reliance upon the adrenal medulla to maintain blood pressure (BP) following damage to sympathetic neurons, and further, the findings are consistent with the possibility that ADR and NA chromaffin cells are separately controlled.

Characterization of methionine-enkephalin release in the rat striatum by in vivo dialysis: Effects of gamma-hydroxybutyrate on cellular and extracellular methionine-enkephalin levels

The opioïd system is implicated in mediating the effects produced upon administration of gamma-hydroxybutyrate. Gamma-hydroxybutyrate occurs endogenously in the mammalian brain, and is most probably involved in the regulation of some basic brain functions, particularly those concerning the dopaminergic nigrostriatal pathway, which is closely linked to the expression of enkephalins in the striatum. In the present study, in vivo microdialysis was used to examine the basic characteristics of methionine-enkephalin (met-enkephalin) release in the striatum of Wistar rats, using a high performance radioimmunoassay. Administration of gamma-hydroxybutyrate to the rats induced a dose-dependent decrease in the extracellular release of met-enkephalin. In parallel, a dose- and time-dependent gamma-hydroxybutyrate-induced accumulation of met-enkephalin in striatum was observed. These two phenomena (tissue accumulation and inhibition of release) were blocked by NCS-382, a gamma-hydroxybutyrate receptor antagonist. The striatal met-enkephalin accumulation does not seem to be exclusively due to the inhibition of its release. Thus, a gamma-hydroxybutyrate mediating effect on met-enkephalin synthesis is suggested, most probably occurring via functional modulation of striatal dopamine synthesis and release. To understand the role of this dopaminergic mechanism, unilateral lesions of the nigrostriatal dopaminergic pathway were carried out. In gamma-hydroxybutyrate-treated rats, striata exhibited a similar increase in met-enkephalin content. In untreated rats, only the lesioned striatum showed an identical increase in met-enkephalin levels. Thus, striatal met-enkephalin accumulation could be attributed to the suppression of the dopaminergic impulse flow, due to gamma-hydroxybutyrate or to the action of 6-hydroxydopamine. In the extracellular spaces (microdialysis experiments), gamma-hydroxybutyrate administration induced identical modifications of met-enkephalin release in lesioned or non-lesioned striata. These modifications could be reproduced by peripheral or striatal administration of sulpiride, a D 2/D 3 antagonist. From a functional point of view, the dopaminergic D 2 receptor blockade or the gamma-hydroxybutyrate-induced inhibition of dopamine release could be considered to induce similar results, with identical consequences on striatal met-enkephalin accumulation and release. These results suggest that gamma-hydroxybutyrate-induced modifications in metenkephalin release, presumably potentiated by 6-hydroxydopamine treatment, act via a functional modification of the nigrostriatal dopaminergic pathway.

Selective effects of retinal dopamine depletion on partial ischemia-induced electroretinographic hyperresponses in rabbits.

The interaction of retinal dopamine depletion and partial ischemia on the a- and b-wave amplitudes and implicit times of the electroretinogram was examined in adult pigmented rabbits. Seven days after 6-hydroxydopamine treatment, which resulted in a depletion of the amine, partial retinal ischemia was induced by raising the intraocular pressure. As expected, moderate elevation of intraocular pressure produced increases in both a- and b-wave amplitudes. Amplitude hyperresponses were significantly reduced in dopamine-depleted retinas. These reductions were more prominent with relatively lower intensities. However, response delays were not shortened but lengthened by 6-hydroxydopamine pretreatment. Together, these results point to a selective role of dopamine in partial retinal ischemia induced by moderate elevation of intraocular pressure in rabbits.

Differential regulation of glutamate decar☐ylase and preproenkephalin mRNA levels in the rat striatum

The mRNA levels encoding for the enzyme glutamate decar☐ylase (GAD67) and the peptide enkephalin were measured in the striatum of adult and 15 day-old rats by in situ hydridization histochemistry and radioautography after neonatal injections of 6-hydroxydopamine or after acute pharmacological blockade of dopamine receptors with haloperidol or sulpiride. In adult rats injected as neonates with 6-hydroxydopamine or treated with the D 1/D 2 dopamine receptors antagonist, haloperidol, an increase in preproenkephalin and GAD67 mRNA levels was measured in the striatum. The D 2 dopamine receptor antagonist, sulpiride, did not change the mRNA levels of either GAD67 or PPE in the striatum. In 15-day-old rats, neonatal 6-hydroxydopamine or haloperidol treatment resulted in increased preproenkephalin but unchanged GAD67 mRNA levels compared to controls. In these 15-day-old rats, however, sulpiride produced an increase in GAD67 but not preproenkephalin mRNA levels. Intrastriatal injections to adult rats of pertussis toxin which uncouples Gi/Go proteins from their receptors resulted in a dramatic increase in preproenkephalin without concomitant change in GAD67 mRNA levels. Altogether, these results show that GAD67 and preproenkephalin mRNA levels are modulated in parallel in adult but not in 15 day-old rats after 6-hydroxydopamine injections or dopaminergic blockade. In keeping with evidence of a co-localization of GAD67 and preproenkephalin mRNAs in some striatal neurons, the results indicate that these two mRNAs can be differentially regulated in the same neurons. In addition, the differential effect of haloperidol, sulpiride or pertussis toxin on GAD67 and preproenkephalin mRNA levels suggests that these two mRNAs are regulated through different dopamine receptor subtypes.

Sympathetic Noradrenergic Fibers as the Source of Immunoreactive Alpha-Neoendorphin and Dynorphin in the Guinea Pig Heart

Dynorphin and alpha-neoendorphin bind to the kappa subtype of opioid receptors and have been shown to inhibit the release of noradrenaline from cardiac sympathetic axons. The purpose of this study was to elucidate the endogenous localization of dynorphin and alpha-neoendorphin in the guinea pig heart. This goal was achieved by double- and triple-labelling immunofluorescence. Dynorphin- and alpha-neoendorphin-immunoreactive nerve fibers were numerous around coronary blood vessels and among cardiomyocytes. They also contained immunoreactivities to the rate-limiting enzyme of catecholamine synthesis tyrosine hydroxylase and to neuropeptide Y. These fibers disappeared in response to chemical sympathectomy (6-hydroxydopamine treatment). In contrast, substance P/calcitonin gene-related peptide-immunoreactive axons of sensory origin did not contain dynorphin and alpha-neoendorphin immunoreactivities and were unaffected by chemical sympathectomy. The findings demonstrate that immunoreactive dynorphin and alpha-neoendorphin are contained in postganglionic sympathetic nerve fibers innervating coronary blood vessels and cardiac muscle. Therefore, the inhibitory effect of these peptides upon noradrenaline release from the sympathetic terminal may well be an autoinhibitory feedback loop.

Adrenergic innervation of the urinary bladder body in the cat with special reference to structure of the detrusor muscle: an immunohistochemical study of noradrenaline and its synthesizing enzymes.

The distribution of adrenergic nerves in the body detrusor muscle of the cat urinary bladder was studied by means of the immunohistochemical identification of noradrenaline (NA) and the NA synthesizing enzymes tyrosine hydroxylase, aromatic L-aminoacid decarboxylase and dopamine beta-hydroxylase. We identified the basic structural organization of the detrusor muscle, which had previously been described as lacking discernible layers. In the lateral wall, both outer longitudinal and inner circular muscle bundles were present, the latter extending in both anterior and posterior directions. The posteriorly running bundles came to lie on the outside of the posterior wall where they enabled recognition of inner longitudinal muscle bundles. Those running anteriorly were dispersed to enter the longitudinal bundles in the anterior wall. NA-immunoreactive nerve fibers in the detrusor muscle of the bladder were found to be similar to those immunoreactive for NA synthesizing enzymes in both distribution and density. In the upper and middle bladder body--including the dome (apex)--immunoreactive nerve fibers were always more abundant in the outer part of the detrusor muscle than in the inner part, regardless of the course of muscle bundles. Even in individual muscle bundles running from the inside to the outer surface, the outer part was more richly innervated by immunoreactive fibers than the inner part. In the bladder dome, a moderate number of immunoreactive nerve fibers preferentially innervated the outer part of the muscle layer. In the lower bladder body, these nerve fibers increased in density in the inner part of the detrusor muscle. There was no sexual difference in density or distribution of nerve fibers. NA- and NA synthesizing enzyme-immunoreactive nerve fibers were markedly decreased in number after 6-hydroxydopamine treatment. No dopamine- or phenylethanolamine-N-methyltransferase-immunoreactive nerve fibers were present in the bladder. The findings of this study indicate that the cat bladder musculature includes longitudinal and circular muscle bundles, both of which are extensively innervated by adrenergic nerves, particularly in the outer part of the bladder.

Dose-related effects of a neonatal 6-OHDA lesion on SKF 38393- and m-chlorophenylpiperazine-induced oral activity responses of rats

Neonatal 6-hydroxydopamine (6-OHDA) treatment of rats is associated with concurrent supersensitization of dopamine (DA) D 1 and serotonin 5-HT 1C receptors, for agonist-induced oral activity. The present study was conducted to determine if graded reduction of striatal DA content and/or graded elevation of striatal 5-HT content by 6-OHDA would alter sensitivity of either receptor type, and thereby influence oral activity responses to DA and 5-HT agonists. At 3 days after birth, groups of rats were pretreated with desipramine (20 mg/kg i.p.), 1 h before administration of a range of doses of 6-OHDA HBr (15, 30, 60, 100, 150 and 200 μg, i.c.v., salt form; half in each lateral ventricle) or the vehicle, saline (0.85%)-ascorbic acid (0.1%). Between 2 and 4 months, a series of challenge doses of SKF 38393 HCl (0.30 to 3.0 mg/kg i.p.) and m-chlorophenylpiperazine 2HCl (0.30 to 6.0 mg/kg i.p.; m-CPP 2HCl) were administered to each group of rats and oral activity was observed. Oral activity was determined for 1 min every 10 min during a 60-min period, starting 10 min after injection of agonist or vehicle. SKF 38393 dose-response curves demonstrated enhanced oral activity responses in rats lesioned neonatally with 150 or 200 μg of 6-OHDA. m-CPP dose-response curves demonstrated enhanced oral activity responses in these 2 groups of rats, as well as those lesioned neonatally with 100 μg of 6-OHDA. Striatal DA content was reduced by > 97% in these 3 groups of rats. Striatal 5-HT content was elevated by > 80% in rats treated neonatally with 150- or 200-μg doses of 6-OHDA, and by 50% in rats treated neonatally with the 100-μg dose of 6-OHDA. Lower doses of 6-OHDA produced less of an effect on striatal DA and 5-HT content. Regression analysis determined that both SKF 38393- and m-CPP-induced oral activities were most closely correlated with the magnitude of change in striatal content of DA. These findings demonstrate that 5-HT agonist responses can be enhanced when DA agonist responses are not enhanced. Also, in neonatal 6-OHDA-lesioned rats the extent of DA depletion vs. the extent of 5-HT elevation seems to be a critical factor in the enhanced behavioral effects of DA and 5-HT agonists.

Ontogenetic SKF 38393 treatments sensitize dopamine D 1 receptors in neonatal 6-OHDA-lesioned rats

Neonatal 6-hydroxydopamine (6-OHDA) treatment of rats is associated with supersensitization of the dopamine (DA) D 1 agonist induction of stereotyped and locomotor behaviors. The present study was conducted to determine whether ontogenetic treatments of these rats with the DA D 1 receptor agonist, SKF 38393, would produce a maximal DA D 1 receptor supersensitivity, as measured by locomotor behavior in adulthood. Rat pups were treated daily with SKF 38393-HCl (3.0 mg/kg per day, i.p.) or saline vehicle for 28 consecutive days from birth. These animals were additionally treated at 3 days after birth with 6-OHDA-HBr (100 μg, in each lateral ventricle, salt form) or its vehicle. Between 6 and 9 weeks locomotor activity or stereotyped behaviors were observed after weekly challenge doses of SKF 38393-HCl (3.0 mg/kg, i.p.). In the neonatal 6-OHDA group, successive SKF 38393 treatments produced progressively greater locomotor activity. In the group of rats treated during postnatal ontogeny with both 6-OHDA and SKF 38393 daily treatments, the first adult challenge dose of SKF 38393 produced an enhanced locomotor response, greater than that seen in other groups ( P < 0.01). Subsequent SKF 38393 treatments of this group produced increasingly greater locomotor responses. SKF 38393-induced stereotyped behavioral effects were greater in the 6-OHDA-lesioned groups, whether or not SKF 38393 was administered ontogenetically. Profound reductions (> 99%) of DA and its metabolites were found in the striatum of neonatal 6-OHDA treated rats, regardless of whether SKF 38393 was co-administered ontogenetically. A marked elevation in striatal 5-HT (> 50%) accompanied the DA depletion in the striatum. These findings indicate that neonatal 6-OHDA treatment produces the expected destruction of striatal DA fibers with associated sprouting of 5-HT fibers, while repeated ontogenetic treatments of these rats with a D 1 agonist produces partial sensitization of the DA D 1 receptors in adulthood.

Age-dependence of a 6-hydroxydopamine lesion on SKF 38393- and m-chlorophenylpiperazine-induced oral activity responses of rats

Neonatal 6-hydroxydopamine (6-OHDA) treatment is associated with destruction of dopamine (DA) fibers and subsequent sprouting of serotonin (5-HT) fibers in the striatum of rats. Enhanced oral activity responses to SKF 38393 and m-chlorophenylpiperazine (ifm-CPP), respective agonists for the DA D 1 receptor complex and 5-HT 2C receptor complex, ensue. To study the ontogenetic nature of this effect, rats were treated at birth, 3 days, 7 days, 10 days or 14 days with 6-OHDA-HBr (200 μg i.c.v.; salt form), following desipramine-HCl pretreatment (20 mg/kg i.p., 1 h; base form). Another group of rats was treated at 35 days and again at 42 days with 6-OHDA-HBr (300 γg i.c.v.), following desipramine-HCl (20 mg/kg i.p., 1 h) and pargyline-HCl (50 mg/kg i.p., 30 min). In rats treated from birth to 10 days, 6-OHDA reduced striatal DA content at 5 months by ≥ 94%. Striatal 5-HT content was elevated by 28% to 51%, but only in rats treated with 6-OHDA at 7 days from birth or earlier. An enhanced oral activity response to SKF 38393-HCl (0.03 to 1.0 mg/kg i.p.) was absent in rats treated 7 days or later, and the change in SKF 38393 effect was correlated with a change in striatal DA content. An enhanced response to m-CPP-2HCl (0.3 to 6.0 mg/kg i.p.) was absent after treatment at 14 or 35 days, when striatal DA content was reduced only 44% to 63% and 5-HT content was not changed. Loss of the enhanced m-CPP response was not directly correlated with the magnitude of change in striatal content of either DA or 5-HT. The findings indicate that SKF 38393 and m-CPP-enhanced oral activity responses are dependent on the age at which 6-OHDA is administered to rats, and that the enhanced response to m-CPP can persist when there is no enhanced response to SKF 38393.

Differential downregulation of beta 2-adrenergic receptors in tissue compartments of rat heart is not altered by sympathetic denervation.

With agonist stimulation, cardiac beta 2-adrenergic receptors (beta 2ARs) are downregulated to a much greater extent than are beta 1ARs. It has been hypothesized that this effect is due to sympathetic innervation inhibiting the downregulation of beta 1ARs. To test this hypothesis, the technique of coverslip autoradiography was used to localize and quantify beta 1AR and beta 2AR subtypes in tissue compartments of the heart in rats subjected to sympathetic denervation by two intravenous injections of 6-hydroxydopamine (50 mg/kg per dose). After denervation, the rats were infused with L-isoproterenol (400 micrograms.kg-1 x h-1 for 7 days) or vehicle (0.001N HCI) by implantation of osmotic minipumps. Sections were incubated with 70 pmol/L of the beta AR antagonist [125I]iodocyanopindolol (ICYP) alone or in the presence of 5 mumol/L DL-propranolol or 5 x 10(-7) mol/L CGP 20712A (a beta 1AR antagonist). Binding of ICYP to sections of rat hearts was saturable and stereoselective and was displaced by beta AR agonists with the rank order of potency expected for beta ARs. There was an 89% reduction in catecholamine concentration in rat ventricles after 1 week of 6-hydroxydopamine treatment, before implantation of the minipumps. Chronic infusion of isoproterenol induced significant downregulation (63% to 74%) of beta 2ARs in atrial and ventricular myocytes, coronary arterioles, and connective tissue but no change in beta 1ARs in these regions in rats with intact sympathetic innervation. Similar changes were seen in denervated animals. There was a marked reduction in beta 2ARs but small insignificant decreases in beta 1ARs, despite the fact that in the denervated animals there was upregulation of beta 1ARs in atrial and ventricular myocytes (approximately 25%). Our study suggests that beta 1ARs in the heart are not significantly downregulated by chronic agonist exposure and that this is unrelated to sympathetic innervation. The underlying mechanism of preferential regulation of beta AR subtypes remains to be elucidated but may be related to differences in the molecular structure between beta 1ARs and beta 2ARs.

Immuno-electron microscopic study of tyrosine hydroxylase in the cat urinary bladder and proximal urethra

The distribution and density of tyrosine hydroxylase (TH)-immunoreactive nerves in the cat urinary bladder and proximal urethra were similar to those of glyoxylic acid fluorescent nerves. Both TH-immunoreactive and fluorescent nerve fibres markedly decreased after 6-hydroxydopamine treatment. Hence, immuno-electron histochemistry of TH was employed to investigate adrenergic termination to the muscle layers of the cat urinary bladder and proximal urethra. A total number of 3728 TH-immunoreactive axon terminals in the lower bladder body, lateral bladder base, trigone, bladder neck and proximal urethra were examined. In the bladder base (the lateral bladder base, trigone and bladder neck) and proximal urethra, most of the TH-immunoreactive axon terminals (57.1–89.2%) lay outside smooth muscle cell fascicles, while most (69.7%) in the bladder body were inside muscle cell fascicles. The proportions of TH-immunoreactive axon terminals accompanying non-TH-immunoreactive axon terminals in the five regions were almost the same (about 70%). The present study demonstrated that the mode of adrenergic termination in the bladder base and proximal urethra notably differed from that in the bladder body, and that approximately 70% of adrenergic axon terminals were associated with non-adrenergic or cholinergic axon terminals in each region.

6-Hydroxydopamine treatment of neonatal rats. I. Effects on the development of the spinal cord noradrenergic system

The spinal cord contains noradrenergic (NA) pathways which descend from cell bodies in the medulla oblongata and pons to terminate at all levels in the spinal gray matter. The present studies sought to determine the patterns of postnatal development of pre- and postsynaptic elements of NA transmission in the spinal cord. Significant presynaptic development is evident at birth as reflected by substantial high-affinity uptake of norepinephrine (NE) into synaptosomes (0.65–0.90 pmol/mg protein). There is a subsequent increase in uptake on postnatal day (PND) 5, followed by a decrease in 5–10 days to essentially adult levels, starting on PND 20 (0.30–0.35 pmol/mg protein). This decrease in NE uptake occurs coincident with increases in the density of postsynaptic α 1 and β adrenergic receptors and also NE-stimulated accumulation of 3′,5′-cyclic adenosine monophosphate (cAMP). Peaks in the development of α 1 receptors (PND 10) and β receptors (PND 20) and NE-stimulated cAMP accumulation (PND 15) were also followed by decreases to adult levels. The neurotoxin 6-hydroxydopamine (6-OHDA) was administered at birth to determine the effects of denervation on the development of the spinal NA systems. At each day following 6-OHDA, synaptosomal uptake of [ 3H]NE was reduced by two-thirds compared with control values. α 1 and β adrenergic receptor binding are uniformly increased along with a parallel increase in NE-stimulated accumulation of cAMP. While uniformly increased over control, the pattern of postnatal increases and decreases in receptors and cAMP accumulation is maintained. These results suggest that developmental signals for the expression of α 1 and β receptors and adenylate cyclase activity in the postsynaptic receptor systems are maintained despite the loss of presynaptic input from descending NA pathways.

6-Hydroxydopamine treatment of neonatal rats. II. Effects on the development of the hindlimb flexor reflex

The flexor reflex (FR) of the hindlimb was measured in rats at various ages between postnatal days (PND) 12–45, in vehicle-treated control rats and in rats treated with 6-hydroxydopamine (6-OHDA) (100 mg/kg, i.p.) on the first and second days after birth. The hindlimb FR was elicited by graded electrical stimulation of the footpad (0.1–2.0 mA) and quantified by the amplitude of the flexion in grams. The half maximal FR response was evoked by 1.2 mA and was 2–3 g in animals at PND 12–17 and 9–10 g in animals on PND 30 and 45. A similar age dependency was evident in the maximum hindlimb FR evoked by 2.0 mA; the maximum FR was 4.7 ± 0.5g on PND 12 and 24 ± 2g on PND 45. In rats treated with 6-OHDA, the strength of the FR was similar to that of the controls up to PND 15. However, the FR was increased 25% by PND 17 and 200% by PND 45 in the 6-OHDA-treated animals, versus controls. Pretreatment with clonidine (100 μg/kg, i.p.), which activates α 1 receptors under the current experimental conditions, did not enhance the FR in control animals. However clonidine pretreatment caused an increase of 400–800% in the FR in the 6-OHDA-treated animals at PND 15 and beyond. Our companion paper demonstrated that as early as PND 5, there was a significant increase ( P < 0.05) in alpha 1 receptors in the spinal cord of 6-OHDA-treated animals, versus the controls 34. The temporal disparity between increased alpha 1 receptors and the increased FR in 6-OHDA-treated animals suggests that factors other than increased α 1 receptors might be involved in the enhancement of the FR.

Amino acid levels and gamma-aminobutyric acidA receptors in rat neostriatum, cortex, and thalamus after neonatal 6-hydroxydopamine lesion.

The amino acid gamma-aminobutyric acid (GABA) is the main inhibitory neurotransmitter in brain, and GABAergic neurons have been proposed to play a major role in basal ganglia physiology. In the neostriatum (caudate putamen), medium-sized aspiny interneurons, as well as neostriatal output neurons that project to several brain regions, use GABA as their neurotransmitter. Dopamine fibers arising from the substantia nigra represent a major input to the neostriatum where, besides their classic neurotransmitter role, they are seemingly involved in the regulation of amino acid neurotransmitter release. To further characterize the nature of some of the amino acid/dopamine interactions, selective dopaminergic deafferentations were produced in neonatal rats (3 days postnatal) by intraventricular administration of the neurotoxin 6-hydroxydopamine (6-OHDA); the noradrenergic neurons were protected by prior administration of desmethylimipramine. After a 3-month survival, levels of catecholamines, indoleamines, and amino acids were determined in cingulate cortex, thalamus, and neostriatum. In addition, GABAA receptors were measured in membrane preparations from these three regions, using the specific agonist [3H]muscimol. In the 6-hydroxydopamine-lesioned rats, levels of dopamine and its metabolites homovanillic acid, 3,4-dihydroxyphenylacetic acid, and 3-methoxytyramine were decreased, as expected, in cortex and neostriatum, but remained unmodified in thalamus. In all three regions, serotonin content was increased; its metabolite, 5-hydroxyindole-3-acetic acid, was also elevated, but only in cortex and neostriatum. The levels of GABA were increased in neostriatum and thalamus, but remained unmodified in cortex. Glycine was increased in all three regions examined. There were also increases of phosphatidylethanolamine and serine in thalamus, and of aspartic acid and alanine in neostriatum. The density of GABAA binding sites was increased in neostriatum, but remained unchanged in cortex and thalamus. The changes in amino acid levels and [3H]muscimol binding sites induced by a neonatal 6-hydroxydopamine treatment differ from those found after similar lesions in adult animals, possibly because of the plastic and synaptic rearrangements that can still occur during early postnatal development. The present results also demonstrate that adaptations occur in response to a dopaminergic deafferentation at an early age and that these exhibit a regional specificity.

Effects of melatonin on pulmonary and coronary vessels are exerted through perivascular nerves

The effects of melatonin on porcine pulmonary and coronary vessels have been studied. Vessels were isolated from normal pigs, placed in a tissue bath, and precontracted with 30 mM KCl. The biophysical responses to cumulative doses of melatonin were then assessed. In the pulmonary artery, melatonin caused a dose-dependent relaxation which was blocked by vasoactive intestinal peptide antagonists or prior 6-hydroxydopamine treatment of the vessels. In the coronary artery, melatonin caused the reverse, a dose-dependent contraction which was blocked by the alpha antagonist prazosin or prior 6-hydroxydopamine treatment. These experiments indicate that melatonin has different mechanisms of action in the coronary and pulmonary circulation, although both seem to depend on the integrity of perivascular 6-hydroxydopamine sensitive nerves. Such differing mechanisms may provide insight into pathophysiological events in the lungs (such as nocturnal asthma) and heart (early morning coronary infarction).

In vivo electrochemical measurements and electrophysiological studies of rat striatum following neonatal 6-hydroxydopamine treatment

The effects of neonatal treatment (one day after birth) with the neurotoxin, 6-hydroxydopamine (75 μg/10 μl intracisternal), were studied in the striatum of normal adult and treated Sprague-Dawley rats. Measurements of monoamine levels in the dorsal striatum and nucleus accumbens, by high-performance liquid chromatography coupled with electrochemical detection, showed that neonatal 6-hydroxydopamine treatment produced a permanent and massive destruction of striatal dopamine. The effects were more pronounced in the dorsal striatum than in the nucleus accumbens. In addition, serotonin levels were elevated in the rat striatum as a consequence of the neonatal treatment. Rapid chronoamperometric recordings of K +-evoked monoamine overflow using Nafion-coated recording electrodes were investigated in both the dorsal and ventral striatum of control and neonatally lesioned rats. The potassium-evoked responses recorded from the dorsal striatum of the 6-hydroxydopamine-treated rats were significantly reduced in amplitude as compared to controls. In addition, the reduction/oxidation current ratios of the responses were more serotonin-like, in contrast to the dopamine-like current ratios measured in the striatum of untreated animals. In ventral striatum, the amplitudes of the K +-evoked responses were not significantly reduced versus control. However, the K +-evoked signals were more serotonin-like in their electrochemical characteristics as compared to controls. In addition to the release studies, extracellular single-unit electrophysiological recordings were performed in normal and neonatally 6-hydroxydopamine treated rats. The spontaneous discharge rate of striatal neurons in the neonatally 6-hydroxydopamine-treated rats was similar to that of control rats. This is in contrast to dopamine lesions in adult animals, where a marked elevation of the discharge rate is observed. Local applications of dopamine and serotonin into the striatum of neonatally 6-hydroxydopamine-treated rats elicited excitations of striatal cells rather than the normal inhibitory effects seen in control animals. Taken together, these data suggest that loss of striatal dopamine terminals at birth leads to both pre- and postsynaptic alterations in monoamine pathways.

Central α-adrenoceptor subtypes involved in the emetic pathway in cats

The intracerebroventricular (i.c.v.) injection of clonidine, xylazine, adrenaline and methoxamine elicited dose-dependent vomiting in cats in that order of potency. The vomiting induced by clonidine, xylazine and adrenaline was antagonized by i.c.v. yohimbine and phentolamine possessing α 2-adrenoceptor-blocking activity, but not by prazosin showing α 1-adrenoceptor-blocking activity. In contrast, methoxamine-induced vomiting was antagonized by prazosin, but not by yohimbine. The vomitting induced by xylazine and adrenaline was not prevented by i.c.v. 6-hydroxydopamine treatment, but was prevented by i.c.v. reserpine treatment. Ablation of the area postrema with some damage to extremely adjacent areas abolished the vomiting induced by each α-adrenoceptor agonist. These results indicate that both central α 1- and α 2-adrenoceptors are involved in the emetic pathway in cats, although α 2-adrenoceptors seem to have the main role. It is also suggested that monoamines, and in particular 5-hydroxytryptamine in the brain, are involved in the regulation of α-adrenoceptor-mediated vomiting.