Bobby Gough

Acute effects of 3,4-methylenedioxymethamphetamine (MDMA) on monoamines in rat caudate

Extracellular levels of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxyindoleacetic acid (5-HIAA) and serotonin (5-HT) were assayed in the caudate of freely moving rats using microdialysis and high performance liquid chromatography with electrochemical detection (HPLC-EC) to detect changes in their release. Dialysates were assayed at 20-minute intervals for four hours after an intraperitoneal (IP) injection of MDMA (10 mg/kg). In a separate study to determine MDMA effects on total caudate levels of the above neurochemicals, animals were injected IP with MDMA (10 mg/kg) and then sacrificed at 20, 60, 120 and 180 minutes after treatment. Brains were quickly removed, and caudate nuclei were dissected for neurochemical analysis using HPLC-EC. MDMA elicited an amphetamine-like increase in DA release, followed by an increase in DA content. DOPAC and HVA were both reduced in homogenate. 5-HT release was also increased, followed by a drop in caudate homogenate levels by three hours. DA extracellular content was 686% of control at 80 minutes; caudate homogenate levels were 122% at 120 minutes. 5-HT extracellular release was 123% at 20 minutes, then decreased thereafter. It is concluded that the acute effect of MDMA on caudate is at least as great on the DA as it is on the 5-HT system.

Effects of a cold environment or age on methamphetamine-induced dopamine release in the caudate putamen of female rats

Extracellular levels of dopamine (DA) and metabolites as well as serotonin [5-hydroxytryptamine (5-HT)] and 5-hydroxyindoleacetic acid (5-HIAA) were determined in the caudate putamen (CPU) of either 6- or 12-month-old female rats using microdialysis and high-performance liquid chromatography with electrochemical detection (HPLC-ED) before, during, and after four consecutive injections (given at 2-h intervals) of methamphetamine (METH). In 6-month-old rats administered 4 x 5 mg/kg METH at an environmental temperature (ET) of 23 degrees C, peak extracellular DA levels (between 50 and 150 rho g/10 microliters) were attained 30-45 min after each dose of METH while dihydroxyphenylacetic acid (DOPAC) decreased steadily after the first doses of METH until it reached a plateau at 50% of control (550-700 pg/10 microliters) levels. Increases in 5-HT levels during METH administrations paralleled DA increases while 5-HIAA decreases paralleled DOPAC decreases. The total CPU DA and 5-HT content of these rats was about 65% of control at 3 days post-METH. Reducing the ET to 4 degrees C during dosing decreased the peak and average DA levels attained during the 4 x 5 mg/kg METH administration to about 50% of that observed at a 23 degrees C ET. Increasing the dose to 4 x 10 mg/kg METH (4 degrees C ET) increased peak and average CPU DA levels to 200% that observed during 4 x 5 mg/kg METH at a 23 degrees C ET. However, no significant decreases in total CPU DA content of any rats dosed with METH at a 4 degrees C ET were observed 3 days post-METH. In 12-month-old rats dosed with 4 x 5 mg/kg METH (23 degrees C ET), the peak and average extracellular DA levels were only 30-60% that of 6-month-old rats. However, the CPU DA content of older rats was significantly decreased both 3 (30% control) and 14 (60% control) days post-METH. In summary, METH toxicity may not be predicted solely by the extracellular levels of DA attained during METH administration; age and ET also greatly influence METH neurotoxicity.

Prenatal dexamethasone or stress but not ACTH or corticosterone alter sexual behavior in male rats

Prenatal maternal stress in rats and mice can demasculinize and feminize the sexual behavior of adult male offspring. Causal mechanisms are unknown, but one attractive hypothesis is that stress activation of maternal adrenal glucocorticoid secretion is the responsible agent. To test this hypothesis, pregnant rats were exposed to a variety of substances which enhance glucocorticoid actions. These included ACTH (20 IU of a gel preparation, SC once daily), corticosterone (CORT; 7 mg/kg SC in oil, three times daily), or dexamethasone (DEX; 0.1 mg/kg, SC once daily). Controls included noninjected dams and a positive stress control group (restraint under bright lights three times daily). All treatments reduced maternal weight gain, DEX most potently. No treatment altered litter size, stillbirths, or sex ratio, but DEX reduced weight at birth, an effect still seen at postnatal day 85. DEX, CORT, and stress reduced male adrenal weight at birth, while DEX and CORT altered sexual differentiation as measured by anogenital distance. Stress impaired adult male sexual performance but not the lordosis quotient following exposure of animals to stud males. DEX affected both measures. No other treatment had any significant effect on sexual behavior. No treatment altered plasma LH levels, either basal or in response to an estrogen challenge in adult gonadectomized males. In adulthood there was no treatment effect on stress reactivity, measured behaviorally or by plasma glucocorticoids. Correlational analysis revealed that weight gain during pregnancy was the single best predictor of subsequent sexual performance. It is concluded that prenatal dexamethasone exposure demasculinizes and feminizes male offspring.(ABSTRACT TRUNCATED AT 250 WORDS)

Nitric oxide regulation of methamphetamine-induced dopamine release in caudate/putamen

A possible role for NO modulation of dopamine (DA) release in the caudate/putamen (CPU) during methamphetamine (METH) exposure was investigated using in vivo microdialysis in rats. Inclusion of the nitric oxide synthase (NOS) inhibitors NG-nitro-L-arginine (NOARG), NG-nitro-L-arginine methyl ester (L-NAME) or D-NAME (less potent inhibitor) in the microdialysis buffer prior to METH minimally affected basal levels of DA, DOPAC or HVA in CPU microdialysate. However, L-NAME and NOARG produced concentration-dependent decreases of up to 64% (100 microM) in CPU DA levels in microdialysate during exposure to four doses of METH (5 mg/kg i.p./2 h), with lesser effects on DOPAC or HVA. Reversal of the NOARG inhibition was produced by inclusion of 500 microM of either L-arginine or L-citrulline in the microdialysate. D-NAME (100 microM) minimally affected levels of DA or metabolites. Paradoxically, inclusion of from 20 to 2 microM of the NOx generators isosorbide dinitrate (ISON) or sodium nitroprusside (SNP) in the microdialysis buffer decreased DA and DOPAC levels in microdialysate during METH exposure. This paradox might result from the concentrations of NOx produced by SNP or ISON being great and not regionally specific resulting in inhibition of DA release and/or synthesis while the NO generated endogenously during METH exposure may have localized and site-specific actions. Alternatively, NOx may inhibit NOS or other enzymes in the NO synthesis pathway, thereby reducing levels of an intermediate (other than NO) which potentiates DA release. In their entirety, our results indicate that NO generation in the CPU may augment the release of DA during METH exposure.

Comparative effects of substituted amphetamines (PMA, MDMA, and METH) on monoamines in rat caudate: a microdialysis study.

Abstract: Paramethoxyamphetamine (PMA) is a methoxylated phenethylamine derivative that has been used illicitly in Australia since 1994. PMA is also becoming popular at rave parties in the United States. PMA raised concern when a series of fatalities resulted after its use in South Australia, where it was marketed as “ecstasy,” which is the colloquial name for MDMA. In the present study, we evaluated the comparative neurotoxicity of substituted amphetamines in rats. Extracellular levels of dopamine (DA), 3,4‐dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), serotonin (5‐HT), and 5‐hydroxyindoleacetic acid (5‐HIAA) were assayed in the caudate of freely moving rats using microdialysis and HPLC‐EC. Dialysates were assayed every 20 minutes for 4 hours after an intraperitoneal (i.p.) injection of PMA (2.5, 5, 10, 20 mg/kg), MDMA (10 and 20 mg/kg), or METH (2.5 mg/kg). METH produced a significant increase in extracellular DA (700%), and significant decreases in extracellular DOPAC and HVA (30% and 50%), with no detectable changes in either 5‐HT or 5‐HIAA. MDMA produced significant increases in DA (700% at 10 mg/kg and 950% at 20 mg/kg) and decreases in DOPAC (15% for both 10 and 20 mg/kg), and HVA (50% at 10 mg/kg and 35% at 20 mg/kg). MDMA also increased 5‐HT (350% at 10, and 575% at 20 mg/kg), and decreased 5‐HIAA to 60% for both dose levels. PMA produced no detectable increases in DA at dose levels of 2.5, 5, or 10 mg/kg, but significantly increased DA (975%) at a dose of 20 mg/kg. However, PMA significantly decreased DOPAC at all dose levels (75% at 2.5; 40% at 5; 30% at 10; 10% at 20 mg/kg), with comparable decreases in HVA at all dose levels. PMA also produced significant increases in 5‐HT at 10 and 20 mg/kg (350% for both dose levels), with no detectable changes in 5‐HT at 2.5 or 5 mg/kg. All dose levels of PMA significantly decreased 5‐HIAA (50 to 70%). These data suggest that PMA, like MDMA and METH, is capable of producing dopaminergic and serotonergic neurotoxicity.

Adrenocortical, β-endorphin and behavioral responses to graded stressors in differentially reared rats ☆

Isolation rearing has long been suspected to alter hormonal and behavioral responses to stress. Two experiments were conducted to test the hypothesis that isolates are more timid or fearful than socially reared rats when exposed to novel test environments. In both, isolate response to 3 graded stressors was compared to that of socially-reared rats. In the first experiment, animals were handled, shocked or not treated prior to testing to produce three levels of conditioned fear. They were then tested on four paradigms previously shown sensitive to conditioned fear: open field activity, emergence latency, auditory startle, and latency to accept food from the experimenter. In the second experiment, rats were given a 0-, 5- or 20-min forced swim, then sacrificed for analysis of plasma corticosterone and pituitary and hypothalamic beta-endorphin. It was found that isolates showed little evidence of enhanced behavioral timidity, although rearing effects were seen on all 4 behavioral measures. Plasma corticosterone levels increased in a graded fashion over the course of the forced swim, but there was no effect of rearing conditions. While there were no effects of rearing or stress on hypothalamic beta-endorphin, pituitary beta-endorphin content was lower in females than in males, and isolate males had lower pituitary endorphin than social males. In summary, these experiments provide no evidence that isolation rearing produces a primary, global increase in fearfulness, but identify several behavioral and hormonal differences associated with differential housing in rats.

Effects of lifelong dietary exposure to genistein or nonylphenol on amphetamine-stimulated striatal dopamine release in male and female rats

Estrogen modulates baseline and amphetamine-stimulated dopamine (DA) release in the adult female rat striatum. The isoflavone found in soybeans, genistein, is a phytoestrogen and may have comparable effects on striatal DA levels. Similarly, the industrial intermediate and potential endocrine disrupter, para-nonylphenol, has estrogen-like effects. Here, Sprague-Dawley rats were continuously exposed to phytoestrogen-free diets containing 0, 100, or 500 ppm genistein (Experiment 1) or 0 or 200, or 750 ppm nonylphenol (Experiment 2) beginning at conception and continuing throughout. To eliminate estrous cycle influences on DA levels, females were ovariectomized at adulthood. As adults, striatal levels of DA and its metabolites [3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA)] were measured in unanesthetized male and female rats via cerebral microdialysis before and for 200 min after an intraperitoneal injection of 2 mg/kg D-amphetamine. Although baseline 5-hydroxyindoleacetic acid (5-HIAA) levels indicated an isolated effect in genistein-treated females, there were no meaningful differences among treatment groups in baseline levels of DA, DOPAC, or HVA. However, dietary exposure to 500 ppm genistein significantly potentiated amphetamine-stimulated DA release in males and a similar trend was apparent, but not statistically significant, in females. Dietary exposure to 200 or 750 ppm nonylphenol had no significant effects in males or females. These results suggest that dietary genistein exposure may act similarly to estradiol in augmenting amphetamine-stimulated DA release.

Methamphetamine-stimulated striatal dopamine release declines rapidly over time following microdialysis probe insertion.

To investigate changes in striatal dopamine release over a series of brief methamphetamine (METH) exposures, METH was pulsed three times at 2-h intervals, with the first exposure occurring 2 h after microdialysis probe insertion. Whether METH was administered directly into the striatum via the microdialysate (20 microM of METH for 10 min), or via peripheral intraperitoneal (i.p.) injection (1 mg/kg METH, i.p.), the dopamine (DA) peak elicited by the third METH exposure was only 50% as large as that elicited by the first exposure, 4 h earlier. This decline in the magnitude of METH-induced DA release probably continued over at least 24 h, since the magnitude of a single peak 26 h after probe implantation was only one-seventh of that at 2 h. This reduction in the response to METH was a function of time post-probe insertion, and not of prior METH exposure. Thus, peak size was the same at 6 h post-implantation in animals which received two prior METH pulses or no prior METH pulses, and in both cases this 6-h peak was substantially lower than that at 2 h post-implantation. Circadian influences were also excluded as a factor, because size of the initial METH-induced DA peak did not vary as a function of time of probe implantation. It is concluded that METH-stimulated striatal DA release declines rapidly over time post-probe insertion. When METH exposures occur repeatedly at short intervals, this decline can mimic, but is not caused by, desensitization or depletion in response to prior METH exposure.

C-type natriuretic peptide (CNP) regulates cocaine-induced dopamine increase and immediate early gene expression in rat brain.

The neuropeptide C‐type natriuretic peptide (CNP) is the primary biologically active natriuretic peptide in brain. Using in situ hybridization, the present report demonstrates that CNP regulates egr‐1, c‐fos and junB immediate early gene expression in rat brain. In the frontal cortex, CNP induced immediate early gene expression whereas it inhibited dose‐dependently the cocaine‐induced early gene expression in the dopaminergic projection fields nucleus accumbens and caudate–putamen. CNP may produce its effect directly on dopaminergic neurons because we found that its receptor, guanylyl cyclase GC‐B, was expressed in the mesencephalon where dopaminergic neurons originate, as well as in their projection fields. The inhibition by CNP of the early gene expression elicited by cocaine in the caudate–putamen is correlated with a CNP‐evoked decrease in cocaine‐induced rise in extracellular dopamine, measured by in vivo microdialysis experiments. The significance of the inhibition of cocaine‐induced dopamine release and early gene induction by the endogenous peptide CNP is demonstrated by data indicating that CNP reduced the cocaine‐induced spontaneous locomotor activation. By inhibiting dopaminergic neuronal activity, CNP represents a potential negative regulator of related behavioural effects of cocaine.

Neurochemical effects of prenatal haloperidol exposure

The neurochemical effects of prenatal exposure to dopamine receptor antagonists are as yet poorly characterized. To further examine this problem, pregnant rats were given daily subcutaneous injections of vehicle, 2.5 or 5.0 mg/kg haloperidol over gestational days 6 through 20. Membrane binding of [3H]SCH-23390 (D1-specific) and [3H]spiroperidol (D2-specific in most brain areas) was measured in four regions of the cerebral dopamine system at postnatal day (PND) 30. Dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) levels were measured in caudate on PND 30 following a d-amphetamine challenge. Prenatal haloperidol exposure reduced [3H]SCH-23390 and [3H]spiroperidol binding in caudate in a dose-dependent manner. [3H]Spiroperidol binding was similarly reduced in nucleus accumbens, but only the low dose (2.5 mg/kg) group showed decreased [3H]SCH-23390 binding in this region. Binding of neither compound was significantly altered in amygdala or frontal cortex. Basal or drug-stimulated levels of caudate DA and DOPAC were unaltered. It is concluded that prenatal haloperidol exposure reduces D1 and D2 binding in some, but not all regions of the forebrain dopamine system.