Chromogranin A inhibits dopamine release from rat striatal slices

Chromogranin A (CGA), a prohormone and a protein component of endocrine and neural secretory granules, neuritic plaques in Alzheimer's disease and Lewy bodies in Parkinson's disease, inhibited the release of dopamine (DA) from perfused rat striatal slices. Dopamine release was stimulated by a pulse of high potassium (40mM) medium introduced at 20 minutes (K1) and 55 minutes (K2) following equilibration. The ratio of K2/K1 was 0.80+/-0.04 in control tissues, but fell significantly to 0.26+/-0.08 when 100nM purified CGA was added prior to the second potassium pulse. This reduction in DA release was equivalent to that seen when calcium was excluded from the buffer (0.19+/-0.05). Pancreastatin, a centrally active peptide product of CGA, had no effect on stimulated DA release (0.77+/-0.06), although it, as well as the other treatments, did reduce basal DA release. It is likely that the parent molecule itself, CGA, or an as yet unidentified product is responsible for inhibition of K-stimulated striatal DA release.     

Protein kinase C inhibitors block amphetamine-mediated dopamine release in rat striatal slices

The stimulant drug amphetamine is postulated to enhance dopamine release through the plasmalemmal dopamine transporter by an exchange diffusion with synaptosomal dopamine. Because protein kinase C has been shown to have an effect on dopamine transporter activity, we examined the effect of protein kinase C inhibitors on endogenous dopamine release stimulated by amphetamine in perfused rat striatal slices. At concentrations of 1 microM, the selective protein kinase C inhibitors chelerythrine, Ro31-8220 and calphostin C nearly completely inhibited endogenous dopamine release elicited by 1 microM amphetamine. The inactive analog bisindoylmaleimide V had no effect. Extracellular Ca++ was not required for the effect of the inhibitors. The importance of vesicular dopamine release was examined by determining inhibitor activity in reserpine-treated rats. Dopamine release elicited by 1 microM amphetamine was not significantly altered in reserpine-treated rats compared with control animals. Ro31-8220 at 1 microM completely blocked amphetamine-induced dopamine release in reserpine-treated rats. Activation of protein kinase C with 250 nM of the phorbol ester 12-O-tetradecanoylphorbol 13-acetate increased dopamine release, and the release was not additive with 1 microM amphetamine. Both chelerythrine and Ro31-8220 at 1 microM increased [3H]dopamine uptake by 17% and 30%, respectively, whereas a brief exposure to 12-O-tetradecanoylphorbol 13-acetate slightly inhibited [3H]dopamine uptake. Our results suggest that amphetamine-mediated dopamine release through the plasmalemmal transporter is highly dependent on protein kinase C activity.     

Modeling voltammetry and microdialysis of striatal extracellular dopamine: the impact of dopamine uptake on extraction and recovery ratios

Numerical modeling was used as a means to examine the relationship between the outcome of in vivo voltammetry and microdialysis experiments and dopamine concentrations in the extracellular fluid of rat striatum. In the case of microdialysis, quantitative interpretation of results demands knowledge of the in vivo values for the extraction and recovery ratios of the probes toward dopamine. Equality of the extraction and recovery ratios is a necessary condition for the direct application of the no-net-flux method as a quantitative technique. Recent results have suggested that the extraction and recovery ratios are not equal, and this interpretation is now supported by theory. A new relationship between extraction and recovery is proposed.     

Influence of selective opiate antagonists on striatal acetylcholine and dopamine release

Ty3 is a retroviruslike element found in Saccharomyces cerevisiae. It encodes GAG3 and GAG3-POL3 polyproteins which are processed into mature proteins found in the Ty3 viruslike particle. In this study, the region encoding a protease that is homologous to retroviral aspartyl proteases was identified and shown to be required for production of mature Ty3 proteins and transposition. The Ty3 protease has the Asp-Ser-Gly consensus sequence found in copia, Ty1, and Rous sarcoma virus proteases, rather than the Asp-Thr-Gly found in most retroviral proteases. The Asp-Ser-Gly consensus is flanked by residues similar to those which flank the active sites of cellular aspartyl proteases. Mutations were made in the Ty3 active-site sequence to examine the role of the protease in Ty3 particle maturation and to test the functional significance of the Ser active-site variant in the consensus sequence. Mutation of the active-site Asp blocked processing of Gag3 and Gag3-Pol3 and allowed identification of a GAG3-POL3 polyprotein. This protein was turned over rapidly in cells expressing the mutant Ty3. Changing the active-site Ser to Thr caused only a modest reduction in the levels of certain Ty3 proteins. Five putative cleavage sites of this protease in Ty3 GAG3 and GAG3-POL3 polyproteins were defined by amino-terminal sequence analysis. The existence of an additional protein(s) of unknown function, encoded downstream of the protease-coding region, was deduced from the positions of these amino termini and the sizes of known Ty3 proteins. Although Ty3 protease cleavage sites do not correspond exactly to known retroviral protease cleavage sites, there are similarities. Residues P3 through P2' in the regions encompassing each of the five sites are uncharged, and no P1 position is occupied by an amino acid with a branched beta carbon.     

Effects of NMDA antagonism on striatal dopamine release in healthy subjects: application of a novel PET approach

We investigated the in vitro erythroid progenitor growth and the effects of sera on normal-marrow CFU-E (colony-forming units - erythroid) growth in 2 patients with renal failure on regular hemodialysis following a prior history of polycythemia vera (PV). PV was diagnosed 3 and 11 years, respectively, before the development of terminal renal failure. One of the patients had entered a spent phase of PV as characterized by diffuse extensive myelofibrosis and anemia; the other also had mild myelofibrosis. The serum erythropoietin (EPO) levels were low or normal on serial measurements by radioimmunoassay. There was no correlation between the hematocrit values and serum EPO levels. EPO-independent erythroid colonies were present in the cultures of bone marrow and peripheral blood cells from both patients after renal failure in the anemic state. With the addition of various concentrations of EPO, the number of erythroid colonies increased as the concentrations of EPO increased which was in accordance with the clinical observation that 1 patient with postpolycythemic myeloid metaplasia partially responded to recombinant human EPO therapy. In the EPO-dependent CFU-E assay, normal-marrow CFU-E numbers supported by 10% of the patient sera were less than those by normal sera. In the absence of EPO in cultures, no erythropoietic activity was found in the patients' sera. Our study on uremic patients with underlying PV showed that the biologic characteristics of autonomous erythroid progenitor growth for PV persisted during the spent phase and after the development of terminal renal failure with anemia. The erythroid progenitors responded to EPO both in vitro and in vivo. Their sera exhibited an inhibiting effect on the growth of normal-marrow erythroid progenitors.     

Dissociation of morphine-induced potentiation of turning and striatal dopamine release by amphetamine in the nigrally-lesioned rat

Morphine has been reported to increase extracellular levels of dopamine in the brain of intact rats and to potentiate turning induced by amphetamine in nigrally-lesioned rats. The present study tested the hypothesis that there is a causal relationship between these two effects of morphine. We tested morphine alone, amphetamine alone, and the combination in separate groups of nigrally-lesioned rats for effects on turning and, by microdialysis, on extracellular dopamine levels. Morphine (3.0 or 10 mg/kg) did not produce significant turning but amphetamine (1.0 mg/kg) did. The lower dose, but not the higher dose, of morphine potentiated amphetamine-induced turning. Amphetamine, but not morphine, produced increases in extracellular dopamine levels. In contrast to what occurred with turning, 10 mg/kg but not 3.0 mg/kg morphine potentiated amphetamine-induced increases in extracellular dopamine levels. These results show that the potentiation of amphetamine-induced turning by morphine in nigrally-lesioned rats is not due to the potentiation of dopamine release in the intact striatum.     

Modulation of [3H]dopamine release by neuropeptide Y in rat striatal slices

Polycythemia vera (PV) is associated with a high incidence of thrombosis. The association of apparent and secondary polycythemia with thrombosis is not clear. It was suggested that activation of the coagulation system contributes to thrombus formation in PV. However, the mechanism of activation is unknown. Monocytes generate a potent tissue factor (TF) upon stimulation with various substances, which is involved in thrombus formation in various disorders. Therefore, we studied the possibility that the factor is involved in the activation of coagulation and thrombus formation also in PV. Unstimulated peripheral blood mononuclear cells (PBMC) from each of the different types of polycythemia expressed weak TF activity (2 U) and antigen (41.4 to 52.9 pg/ml), which were similar to normal controls. Following stimulation with endotoxin, PBMC from normal controls and from apparent and secondary polycythemia showed a 3.9- to 4.5-fold increase in TF, while cells from PV showed a 21-fold increase (P<0.001). Similar levels were generated by PBMC after treatment of PV and at the spent phase. TF was generated by monocytes but not by lymphocytes. Plasma prothrombin fragment1+2 (F1+2) levels, assayed at the same time, were significantly higher in PV (2.46 nm) compared to normals and apparent and secondary polycythemia (0.22 to 0.32 nm), and were in a significant correlation with monocyte TF activity and antigen levels (r = 0.77, 0.87). The high levels of F1+2 confirm that the coagulation system is activated in PV. The increased capacity of monocytes to generate TF may be responsible for the activation of the coagulation system and thrombus formation. The hypercoagulability state that is induced by this mechanism suggests that long-life oral anticoagulation should be considered once thrombosis has been developed in PV.     

Characteristics of the NMDA receptor modulating hypoxia/hypoglycaemia-induced rat striatal dopamine release in vitro

We investigated the functional characteristics of the NMDA receptor that modulates hypoxia/hypoglycaemia-induced striatal dopamine release. Dopamine release was detected by fast cyclic voltammetry in rat neostriatal slices. Four variables were measured: T(on) -- time from initiation of hypoxia/hypoglycaemia to the onset of dopamine release, Tpk -- time from onset to maximum, deltaDA/delta(t) -- rate of dopamine release and DAmax -- maximum extracellular dopamine concentration. In controls, T(on) = 164.9 +/- 1.7 s, Tpk = 20.9 +/- 0.9 s, deltaDA/delta(t) = 5.31 +/- 0.44 microM/s and DAmax = 79.1 +/- 2.5 microM (means +/- S.E.M., n = 203). Cis-4-(phosphonomethyl)piperidine-2-carboxylic acid (CGS 19755, 20 microM) lengthened, while N-methyl-D-aspartate (NMDA) (100 microM) shortened T(on). (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,1 0-imine hydrogen maleate (MK 801, 1 and 10 microM) and dextromethorphan (10 and 100 microM) increased Tpk and decreased DAmax. Neither glycine (100 microM), 7-chlorokynurenic acid (50 microM) nor 5-nitro-6,7-dichloro-1,4-dihydroquinoxaline-2,3-dione (ACEA 1021, 100 microM) had any effect although 7-chlorokynurenic acid blocked the effect of NMDA. Increasing [Mg2+] from 1.3 to 3.7 mM, increased Tpk and decreased deltaDA/delta(t). Dithiothreitol (1 mM) accelerated T(on) while 5.5-dithio-bis-(2-nitrobenzoic acid) (1 mM) delayed T(on). Neither drug affected Tpk, DAmax or deltaDA/delta(t). Neither spermidine (100 microM) nor arcaine (100 microM) affected T(on), Tpk or deltaDA/delta(t) although arcaine decreased DAmax. In conclusion, hypoxia/hypoglycaemia-induced dopamine release was influenced by an NMDA receptor although modulation of the glycine recognition site of the receptor was ineffective, as were agents acting at polyamine modulatory zones. These findings highlight differences between recombinant and native NMDA receptors and suggest caution in extrapolating molecular biology to functional studies.     

Effects of oral exposure to mining waste on in vivo dopamine release from rat striatum

Several single components of mining waste (arsenic, manganese, lead, cadmium) to which humans are exposed at the mining area of Villa de la Paz, Mexico, are known to provoke alterations of striatal dopaminergic parameters. In this study we used an animal model to examine neurochemical changes resulting from exposure to a metal mixture. We used microdialysis to compare in vivo dopamine release from adult rats subchronically exposed to a mining waste by oral route with those from a control group and from a sodium arsenite group (25 mg/kg/day). We found that arsenic and manganese do accumulate in rat brain after 2 weeks of oral exposure. The mining waste group showed significantly decreased basal levels of dihydroxyphenylacetic acid (DOPAC; 66.7 +/- 7.53 pg/ microl) when compared to a control group (113.7 +/- 14.3 pg/ microl). Although basal dopamine release rates were comparable among groups, when the system was challenged with a long-standing depolarization through high-potassium perfusion, animals exposed to mining waste were not able to sustain an increased dopamine release in response to depolarization (mining waste group 5.5 +/- 0.5 pg/ microl versus control group 21.7 +/- 5.8 pg/ microl). Also, DOPAC and homovanillic acid levels were significantly lower in exposed animals than in controls during stimulation with high potassium. The arsenite group showed a similar tendency to that from the mining waste group. In vivo microdialysis provides relevant data about the effects of a chemical mixture. Our results indicate that this mining waste may represent a health risk for the exposed population.     

Differential inhibition by alpha-conotoxin-MII of the nicotinic stimulation of [3H]dopamine release from rat striatal synaptosomes and slices

The presynaptic nicotinic modulation of dopamine release from striatal nerve terminals is well established, but the subtype(s) of neuronal nicotinic acetylcholine receptor (nAChR) underlying this response has not been identified. Recently, alpha-conotoxin-MII has been reported to inhibit potently and selectively the rat alpha3beta2 combination of nAChR subunits. Here we have synthesised the peptide, confirmed its specificity, and examined its effect on the (+/-)-anatoxin-a-evoked release of [3H]dopamine from rat striatal synaptosomes and slices. Alpha-conotoxin-MII (112 nM) completely blocked acetylcholine-evoked currents of alpha3beta2 nAChRs expressed in Xenopus oocytes (IC50 = 8.0 +/- 1.1 nM). Pairwise combinations of other nicotinic subunits were not blocked by 112 nM alpha-conotoxin-MII. On perfused striatal synaptosomes and slices, alpha-conotoxin-MII dose-dependently inhibited [3H]dopamine release evoked by 1 microM (+/-)-anatoxin-a with IC50 values of 24.3 +/- 2.9 and 17.3 +/- 0.1 nM, respectively. The dose-response curve was shifted to the right with increasing agonist concentrations. However, the maximal inhibition of responses achieved by alpha-conotoxin-MII (112 nM) was 44.9 +/- 5.4% for synaptosomes and 25.0 +/- 4.1% for slices, compared with an inhibition by 10 microM mecamylamine of 77.9 +/- 3.7 and 88.0 +/- 2.1%, respectively. These results suggest the presence of presynaptic alpha3beta2-like nAChRs on striatal dopaminergic terminals, but the incomplete block of (+/-)-anatoxin-a-evoked [3H]dopamine release by alpha-conotoxin-MII also supports the participation of nAChRs composed of other subunits. The lower inhibition found in slices is consistent with an additional indirect nicotinic stimulation of dopamine release via an alpha-conotoxin-MII-insensitive nAChR.     

Differential effects of reserpine and tetrabenazine on rat striatal synaptosomal dopamine biosynthesis and synaptosomal dopamine pools

Rat striatal dopamine (DA) levels and synaptosomal DA synthesis were determined after the administration of the catecholamine-depleting agents reserpine (RES) and tetrabenazine (TBZ). Striatal synaptosomal DA synthesis remained unchanged from control levels after Res administration over a wide range of doses (0.5-5 mg/kg) and times (up to 48 hour). In contrast, after TBZ administration, DA synthesis rapidly increased to values greater than 200% of control values, then returned to control levels. The changes in DA synthesis inversely paralleled the depletion of and recovery of striatal DA levels. The increased levels of DA synthesis did not appear to originate with alterations either in the kinetic properties of tyrosine hydroxylase or in the availability of exogenous or endogenous tyrosine. To assess the contribution of synaptosomal DA pools to the regulation of DA synthesis in tissue preparations from RES-or TBZ-pretreated animals, synaptosomal DA synthesis was assessed in the presence of DA releasing agents and compared with analogous experimental manipulations on tissue preparations from animals pretreated with alpha-methyltyrosine or the monoamine oxidase inhibitor, clorgyline. The data are consistent with a differential in vivo interaction of RES and TBZ with a nerve ending pool of DA which participates in end-product inhibition of tyrosine hydroxylase.     

Effect of amygdaloid kindling on [3H]dopamine and [14C]acetylcholine release from rat prefrontal cortex and striatal slices

The involvement of the dopaminergic (DA) systems in the control of limbic kindled seizures is ill defined. The effects of kindling on DA activity may have been overlooked in the past, because of its subtle unilateral occurrence and/or the variance of the endogenous imbalance of DA activity in normal animals. In the present study rats were screened for their endogenous DA imbalance using amphetamine-induced rotational behaviour. Electrical or sham kindling was applied in the hemisphere with the higher endogenous DA activity. Sections of the bilateral prefrontal cortex and dorsal and ventral striatum were dissected either 2 hours or 21 days after the final seizure and the electrically stimulated release of [3H]DA and [14C]acetylcholine (ACh) determined. Release was also measured in the presence of quinpirole or sulpiride to assess the activity of pre- and postsynaptic DA D2-receptors. Long-term effects of kindling consisted of facilitation of ACh release in the ventral striatum contralateral to the kindled amygdala and bilateral depression of DA release in the prefrontal cortex. Kindling therefore produced area specific changes in neurotransmitter systems giving rise to increased pro-convulsive cholinergic activity in the ventral striatum and decreased anti-convulsive dopaminergic activity in the prefrontal cortex.     

Effects of Ca2+ channel antagonists on striatal dopamine and DOPA release, studied by in vivo microdialysis

The chaetotaxy of argentophilic structures on three species of the monogenean genus Gyrodactylus was investigated in an attempt to distinguish species of this genus. Maps were prepared for Gyrodactylus salaris from Scandinavia and compared with two native species of Gyrodactylus parasitising salmonids in Britain, namely Gyrodactylus derjavini and Gyrodactylus truttae. The maps were subsequently refined and analysed for zones of homology and differentiation. The results demonstrate that G. salaris can be readily distinguished by this technique, which is, therefore, of great potential value in the identification of this notifiable pathogen. The key aggregations of sensilla discriminating G. salaris are, ventrally, the antero-ventral set, the medio-lateral set and the postero-lateral set, and, dorsally, the postero-dorsal set.     

The effect of midazolam and propofol on interleukin-8 from human polymorphonuclear leukocytes

Anesthetics and sedatives contribute to postoperative immunosuppression. Interleukin-8 (IL-8) is a chemotactic and activating factor that mediates neutrophil adhesion and margination and is essential for host defense. We investigated the effect of anesthetics on isolated human polymorphonuclear leukocyte production of IL-8. Healthy human polymorphonuclear leukocytes were isolated using a single-step density gradient and stimulated with lipopolysaccharide in the presence of varying concentrations of propofol or midazolam for up to 20 h. IL-8 was measured in both culture supernatants and cell lysates using enzyme immunoassay, and IL-8 mRNA in cells was measured using Northern blotting and phosphorimaging. Data were analyzed using Kruskal-Wallis analysis of variance or the Mann-Whitney U-test as appropriate. Lipopolysaccharide increased extracellular accumulation of interleukin-8, which was suppressed by both propofol (P = 0.025) and midazolam (P = 0.028). However, intracellular IL-8 increased with exposure to lipopolysaccharide (P = 0.028) and remained increased with both anesthetics. Northern blot analysis also revealed increased IL-8 mRNA levels in the presence of both midazolam and propofol, which was confirmed by molecular imaging. These data strongly suggest that the anesthetics modulate transport or secretion of IL-8 protein from the cell. Suppression of IL-8 by anesthetics and sedatives may predispose postoperative and intensive care patients to infection. Implications: Anesthesia causes immune suppression and alters neutrophil function. We investigated the effect of propofol and midazolam on interleukin-8, a neutrophil chemotactic agent in human neutrophils. Both anesthetics decreased extracellular interleukin-8 accumulation, but intracellular levels and mRNA remained high. This suggests that propofol and midazolam alter interleukin-8 secretion from cells.     

Ontogeny of striatal dopamine release in rats after acute administration of methamphetamine

In the present study, we examined the effects of acute MAP administration on striatal extracellular levels of dopamine (DA) and its metabolites in groups of rats on postnatal days (PNDs) 14, 21, 28, and 56. A single injection of 4 mg/kg MAP (IP) induced increase in extracellular DA and decrease in extracellular 3,4-dihydroxyphenylacetic acid (DOPAC) in the striatal perfusates of rats on all PNDs examined. The magnitude of increase in DA concentrations at 20 min after the MAP injection was significantly smaller on PND 14 than PNDs 21, 28, and 56, whereas the magnitude of decrease in DOPAC concentrations after the MAP injection was significantly smaller on PND 14 than PNDs 21, 28, and 56. After the MAP injection, homovanillic acid levels decreased on PNDs 21, 28, and 56, but increased on PND 14. These results suggest that rats on PND 14 differ from those thereafter in MAP-induced DA release and changes in its metabolites, and that such developmental effect on MAP-induced DA release may be involved in the ontogeny of MAP-induced behavioral sensitization.     

Endothelin induces dopamine release from rat striatum via endothelin-B receptors

The aim of the present study was to determine whether local administration of endothelin induces the release of dopamine in the rat striatum and to characterize and localize endothelin receptors in this brain region. Local injection of endothelin-1 (10 pmol) into the ventral striatum of urethane-anaesthetized rats caused an increase of 8 microM in the extracellular concentration of dopamine as measured by in vivo chronoamperometry. The peak increase in dopamine concentration occurred within 5 min of endothelin injection. Injection of the selective endothelin-B receptor agonist [Ala1.3,11.15]endothelin-1 (10 pmol) also caused an increase in extracellular dopamine concentration, suggesting that endothelin is acting at the endothelin-B receptor to elicit its effect. In rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal pathway, the response to local injection of endothelin-1 (10 pmol) was significantly inhibited on the lesioned side as compared to the non-lesioned side. In contrast, pretreatment of the rats with the N-methyl-D-aspartate receptor antagonist dizocilpine maleate (5 mg/kg, i.p.) or the nitric oxide synthase inhibitor NG-nitro-L-arginine (3 mg/kg, i.p.) did not alter the endothelin-induced release of dopamine. In binding studies, addition of endothelin-1 displaced [125I]endothelin-1 with a Ki of 220 pM. The endothelin-B receptor antagonist BQ788 displaced [125I]endothelin-1 with a Ki of 120 nM, whereas the endothelin-A receptor antagonist BQ123 produced only a 25% displacement at 10 microM, suggesting that endothelin receptors in the striatum are of the endothelin-B subtype. In rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal dopamine system, [125I]endothelin-1 binding was reduced by 53% in lesioned striatum compared to non-lesioned striatum, with no difference in the Kd. These data provide evidence that endothelin acts on a homogeneous population of endothelin-B receptors within the striatum to cause the release of dopamine and that a significant proportion of these receptors is located on dopaminergic neurons.     

Effects of the combinations propofol/alfentanil and midazolam/fentanyl on blood pressure and contact phase system during coronary surgery

Perioperative haemodynamic changes leading to severe circulatory problems during open-heart surgery still represent dreaded complications. The aim of this study was to examine the relationship between the use of applied anaesthetic agents and alterations of the contact phase of the intrinsic blood-clotting system, as changes within the kallikrein-kinin system can lead to a fall in blood pressure. In a randomized study, parameters of the kallikrein-kinin system, coagulation and fibrinolysis were determined for 36 patients with aortocoronary bypass operations. The patients had been given either midazolam/fentanyl or propofol/alfentanil to maintain anaesthesia. Perioperative blood pressure values were registered at seven fixed points. The measured values of the factor XIIa-like activity and the kallikrein-like activity suggested a higher activation of the contact phase, when propofol/alfentanil was given. From the start of the extracorporeal circulation (ECC) to the end of the operation, the kallikrein-like activities in the propofol/alfentanil group were significantly higher than those of the midazolam/fentanyl group. Also, the results of the kallikrein inhibition capacity and the indicators of fibrinolysis (t-PA and D-dimers) indicate a stronger activation of the contact phase--at least at the beginning of recirculation--and as a result of it, a stronger fibrinolysis within the propofol/alfentanil group. In addition, the hypotensive side-effects differed significantly between the two groups. Patients receiving propofol/alfentanil needed the triple amount of antihypotonicum to maintain the mean arterial blood pressure above 75 mmHg. With the results of this study, a correlation between the application of propofol/alfentanil, contact phase activation, with activation of the kallikrein-kinin-bradykinin system and the observed hypotension, can be presumed.     

Effect of morphine on striatal dopamine metabolism and ascorbic and uric acid release in freely moving rats

Recent ex vivo findings have shown that morphine increases dopamine (DA) and xanthine oxidative metabolism and ascorbic acid (AA) oxidation in the rat striatum. In the present study, we evaluated the effects of subcutaneous daily morphine (20 mg/kg) administration on DA, dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), AA and uric acid in the striatum of freely moving rats using microdialysis. Dialysates were assayed by high performance liquid chromatography with electrochemical detection. On the first day, morphine administration caused a significant increase in extracellular DA, DOPAC, HVA, AA and uric acid concentrations over a 3 h period after morphine. In all treated rats (n = 7), individual concentrations of DOPAC + HVA were directly correlated with individual AA and uric acid concentrations. Last morphine administration on the 4th day increased DOPAC, HVA, AA and uric acid concentrations but failed to increase those of DA. Individual DOPAC + HVA concentrations were still directly correlated with individual AA and uric acid concentrations. These results suggest that systemic morphine increases both striatal DA release and DA and xanthine oxidative metabolism. Only the former effect undergoes tolerance. The increase in DA oxidative metabolism is highly correlated with that of xanthine. The subsequent enhancement in reactive oxygen species production may account for the increase in extracellular AA.     

Involvement of cholinergic presynaptic receptors of nicotinic and muscarinic types in the control of the spontaneous release of dopamine from striatal dopaminergic terminals in the rat

Rat striatal slices (two) were superfused continuously with L-3,5-3H tyrosine and 3H-dopamine (3H-DA) release was estimated in serial fractions of superfusates. The spontaneous release of 3H-DA was reduced by about 50% when slices were superfused with a calcium-free medium containing ethylene glycol bis (beta-aminoethyl ether)- N,N'-tetraacetic acid (EGTA) (10(-4) M) or with a medium containg tetrodotoxin (5 x 10(-7) M). These effects were not related to a change in 3H-DA synthesis since the rate of L-3,5-3H tyrosine hydroxylation, as measured by 3H-H2O formation was not significantly reduced. Acetylcholine (ACh) (10(-5) M) stimulated the release of 3H-DA (about 100%). This effect was abolished in the absence of calcium; it was partially blocked by pempidine (10(-5) M), atropine (10(-6) M) or scopolamine (10(-5), 10(-6) M). Oxotremorine (10(-5) M) enhanced 3H-DA release but to a lesser extent (60%) than ACh (10(-5) M); its action was completely blocked by atropine (10(-6) M) and unaffected by pempidine (10(-5) M). The ACh- (10(-5) M) and oxotremorine- (10(-5) M) stimulatine effects on 3H-DA spontaneous release were still detected in slices superfused in the presence of tetrodotoxin (5 x 10(-7) M). In the presence of the neurotoxin, the effect of ACh (10(-5) M) was significantly reduced by pempidine (10(-5) M) and the effect of oxotremorine (10(-5) M) was blocked by atropine (10(-6) M). These results suggest the presence of cholinergic presynaptic receptors of the nicotinic and muscarinic types on striatal dopaminergic terminals.