Bromocriptine inhibits incorporation of [H]thymidine into rat pituitary tumor cells

The effects of bromocriptine on GH₃ pituitary tumor cell [³H]thymidine incorporation were studied. Cells were grown in the presence of bromocriptine, then exposed to a short-term pulse of [³H]thymidine in serum-free medium containing deoxycytidine (10 μM) to prevent deoxythymidine triphosphate (dTTP) pooling. After 48 h exposure to bromocriptine, basal prolactin (PRL)-secretion during 45 min was inhibited by 50% by 10 μM bromocriptine and thyroid releasing hormone-induced PRL stimulation was suppressed. Incorporation of radiolabelled thymidine into acid-precipitable DNA increased progressively from 15 to 60 min and was abolished by simultaneous incubation with excess unlabelled thymidine (100 μM). Bromocriptine (10 μM) inhibited incorporation of 5–50 μM [³H]thymidine, but this was not reversed by simultaneous incubation with metoclopramide (10 μM). Aminopterin, an inhibitor of endogenous de novo DNA synthesis, stimulated [³H]thymidine incorporation twofold and this increased DNA salvage pathway activity was also blocked by bromocriptine. As incorporation of [³H]thymidine into acid-soluble cell neucleotides was also inhibited by bromocriptine, the data suggest that in these cells the drug inhibits thymidine kinase activity, a salvage pathway of DNA synthesis.     

The effect of ibogaine on κ-opioid- and 5-HT3-induced changes in stimulation-evoked dopamine release in vitro from striatum of C57BL/6BY mice

Ibogaine is an kndole alkaloid that has bean suggested to have potential efficacy for internrpdng dependermy on stimulant drugs. The κ-opiold and serotonkr 5-HT₃ systems may be involved in the action of Ibogaine, related to their modulation of dopaminergic transmission. The κ-opioid agonist U 62066 attenuated the in vitrostimulation-evoked efflux of tritium label from striatal tissue prelabeled with (₃H)dopamine. In mice protreated with Ibogaine·HCl (40 mg/kg IP given 2 h prior or 2 × 40 mg/kg and animals killed 18 h later), the Inhibitory effect of U 62066 on stimulation-evoked release of tritium was eliminated. The 5-HT₃ agonist phernylbiguanide had a biphasic effect on stimulation-evoked release of tritium; at 10⁻⁶ M phenylbiguanide, stimulation-evoked release was attenuated. At 10⁻⁵ M the basal outflow of tritium was increased. Ibogaine pretreatment had no effect on basal or stimulation-evokedrelease in the presence of 10⁻⁶ M phenylbiguanide, but increased the stimulation-evoked outflow of bftkan in the presence of 10⁻⁵ M phenylbiguanide. Cocaine (10⁻⁶ M), a dopanlne uptake blocker, increased the electrically-evoked release of dopamine; ibogaine pretreatment did not affect the enhanced electrically-induced release of (³H]dopamine by in vitro cocalne. The effects of ibogaine on the κ-opioid and 5-HT₃ receptors, located presynaptically on stristal dopamine terminals, modulating dopamine release may partly underlie its putative antiaddictive properties.     

Not only dopamine D2 receptors involved in Peony-Glycyrrhiza Decoction,an herbal preparation against antipsychotic-associated hyperprolactinemia

Clinical studies have demonstrated the effectiveness of an herbal preparation called Peony-Glycyrrhiza Decoction (PGD) in alleviating antipsychotic-induced hyperprolactinemia (hyperPRL). In the present study, we further examined the pharmacological action of PGD on prolactin (PRL) secretion using in vitro and in vivo models, with specific attention to the role of dopaminergic mediators and other sex hormones. Treatment with PGD at 1–5 mg/ml significantly suppressed PRL secretion and synthesis in MMQ cells, a model of hyperPRL derived from pituitary adenoma cells. The suppressive effects were completely abolished by pretreatment with 10 μM haloperidol, a dopamine D₂ receptor antagonist. Consistent with a D₂-action, PGD did not affect PRL in rat pituitary lactotropic tumor-derived GH3 cells that lack the D₂ receptor expression but significantly increased the expression of D₂ receptors and dopamine transporters (DAT) in PC12 cells. In a rat model of hyperPRL, produced by repeated injection of the dopamine blocker metoclopramide (MCP), chronic PGD (2.5–10 g/kg daily) significantly reduced elevated serum PRL. The reduction in magnitude was similar to that elicited by bromocriptine (BMT), a dopamine D₂ receptor agonist currently used for treatment of hyperPRL. Neither PGD nor BMT altered serum estradiol, but PGD reversed decreased serum progesterone to control level, whereas BMT did not. These results indicate that the anti-hyperPRL effects of PGD are associated not only with D₂ receptor and DAT modulation, but also with a normalization of other sex hormone dysfunction. This experimental evidence supports clinical use of PGD as an effective treatment of antipsychotic-induced hyperPRL.     

Nucleus A10 dopaminergic neurons in inbred mouse strains: Firing rate and autoreceptor sensitivity are independent of the number of cells in the nucleus

Inbred mouse strains have different numbers of midbrain dopaminergic neurons; for example, BALB/cJ mice have 20–25% more neurons than CBA/J mice. As the number of cells decrease, for example in Parkinson's disease and in animals with midbrain dopaminergic cell lesions, the activity of their remaining cells increases. The purpose of the present experiment was to determine whether the functional properties of dopaminergic neurons in the ventral tegmental area (nucleus A10) differ in inbred mouse strains which possess different numbers of cells. The firing rate and autoreceptor sensitivity of A10 dopaminergic cells were examined in the in vitro slice preparation in BALB/cJ, C3H/HeJ, CBA/J, and DBA/2J mouse strains. It was observed that the autoreceptors on mouse dopaminergic neurons exhibit pharmacological properties of dopamine autoreceptors; activation of the autoreceptor produced a marked inhibition (50–70%) in cell firing rate by quinpirole (10⁻⁸ M), LY-141865 (10⁻⁷ M), (+)-3-(3-hydroxyphenyl)-N-n-propyl-piperidine (10⁻⁶ M), propyl-norapomorphine (10⁻⁵ M) and dopamine (10⁻⁴ M), and this inhibition was blocked or reversed by specific dopamine D2 receptor antagonists [(−) sulphide and spiroperidol, 10⁻⁶ M]. The baseline firing rates of the A10 cells did not differ among the four inbred strains [range 2.5 ± 0.2 (C3H/HeJ)−3.4 ± 0.3 (CBA/J) spikes/s ±SEM], and there was no significant difference in autoreceptor sensitivity among the mouse strains as assessed either by superfused dopamine (inhibitory dose 50% ≈150 μM), or by superfused quinpirole (inhibitory dose 50% ≈10 nM). These data indicate that differences in A10 cell numbers of 30% do not significantly influence the baseline firing rate or autoreceptor sensitivity of the cells.     

Inhibition of tyrosine hydroxylation in tissue slices of the rat striatum by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a nigrostriatal neurotoxin in humans and primates, at 10⁻⁵ M inhibited hydroxylation of tyrosine to 3,4-dihydrophenylalanine (DOPA), the rate-limiting step of dopamine synthesis, in tissue slices of the striatum and nucleus accumbens of the rat. Nomifensine, an inhibitor of dopamine uptake, reversed the inhibition but sulpiride, a dopamine receptor antagonist, did not affect the inhibition. MPTP at 10⁻⁵ M inhibited neither the purified tyrosine hydroxylase nor dihydropteridine reductase in vitro. The level of total biopterin did not change significantly, but the tetrahydrobiopterin level was decreased in the striatal slices incubated in the presence of MPTP. These results suggest that MPTP inhibits dopamine synthesis in situ at the tyrosine hydroxylase step probably through inhibition of dihydropteridine reductase.     

Regulation of Dopamine D1 and D2 Receptors on Striatal Acetylcholine Release in Rats

The effects of dopamine (DA) D₁ and D₂ receptors on striatal acetylcholine (ACh) releases were investigated by in vivo microdialysis. All drugs were applied via dialysis membrane directly to the striatum. The levels of ACh release were increased by 10⁻⁴ M SKF38393, a D₁ receptor agonist. Although 10⁻⁴ M SCH23390, a D₁ receptor antagonist, exhibited an increase in the levels of ACh release, the agonist (10⁻⁴ M) induced-increase in the levels of ACh release was suppressed by coperfusion of the antagonist (10⁻⁴ M). In contrast, the levels of ACh release were decreased by the D₂ receptor agonist, N-434, in a dose-dependent manner (10⁻⁵ M to 10⁻⁷ M) and increased by the D₂ receptor antagonist, sulpiride, in a dose-dependent manner (10⁻⁵ M to 10⁻⁷ M). The agonist (10⁻⁵ M) induced-decrease in the levels of ACh release was suppressed by coperfusion of the antagonist (10⁻⁶ M). Coperfusion of D₁ (10⁻⁴ M) and D₂ (10⁻⁵ M) agonists blocked both effects of respective drug alone. In order to clarify the effect of endogenous DA, two drugs with different mechanisms for enhancing DA concentration in the synaptic cleft, the DA release-inducer methamphetamine, and the DA uptake inhibitor nomifensine were perfused separately. Both (10⁻⁴ M to 10⁻⁶ M) produced a dose- and a time-dependent decrease in the levels of ACh release. Significant higher levels of ACh release were observed in the striatum of the 6-hydroxydopamine (8

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)-treated rats with significant depletion of striatal DA content. These results suggest that in striatal DA-ACh interaction ACh release, as cholinergic interneuron's activity, is tonically inhibited via the D₂ receptor, mainly by dopaminergic input, and the D₁ receptor probably modifies the effect of the D₂ receptor indirectly.     

Local and distal effects induced by unilateral striatal application of opiates in the absence or in the presence of naloxone on the release of dopamine in both caudate nuclei and substantiae nigrae of the cat

Halothane-anesthetized cats implanted with push-pull cannulae in both caudate nuclei (CN) and substantiae nigrae (SN) were used to study the effects of naloxone and various opiates when applied into the left CN on the release of newly synthetized tritiated dopamine (DA) from nerve terminals and dendrites of the two nigro-striatal dopaminergic pathways. In all cases, the drugs (naloxone, opiates alone or in the presence of naloxone) were applied for 30 min into the left CN. When applied alone, naloxone (10⁻⁶ M) induced a delayed reduction in tritiated DA release both in the ipsilateral and contralateral CN. These effects were seen after removal of the drug from the superfusion fluid. Complementary experiments made with tritiated naloxone (10⁻⁶ M) revealed that the contralateral effect on DA release was not due to a diffusion of the opiate antagonist from its application site. Locally,d-Ala₂, Met-enkephalinamide (d-Ala₂, Met-Enk, 10⁻⁶ M) and the potent δ agonist Tyr-d-Ser-Gly-Phe-Leu-Thr (DSThr, 5 × 10⁻⁸ M) induced a biphasic increase in tritiated DA release. The local changes in tritiated DA release evoked by morphine (10⁻⁶ M) and μ agonists such as Tyr-d-Ala-Gly-NH-C₆H₁₃ (10⁻⁸ M) and fentanyl (10⁻⁸ M) differed from those of δ agonists and furthermore differed from each other. For instance, morphine induced a delayed increase in tritiated DA release whereas a biphasic increase followed by a delayed inhibition occurred with fentanyl. Among all the opiates testedd-Ala₂-Met-Enk was the only one which elicited a distal effect, that is a reduction of tritiated DA release in the ipsilateral SN. Marked differences in these opiates' effects on tritiated DA release occurred both locally and in distal structures when opiates were applied simultaneously with naloxone (10⁻⁶ M). Locally, the changes induced by μ agonists were particularly altered since during morphine's application with naloxone a reduction of tritiated DA release occurred. In addition, the opiate antagonist prevented the second increase and the delayed inhibition of tritiated DA release evoked by fentanyl (10⁻⁸ M). Interestingly, the combined application of naloxone with eitherd-Ala₂,Met-Enk (10⁻⁶ M) DSThr (5 × 10⁻⁸ M) or morphine (10⁻⁶ M) resulted in the appearance of changes in tritiated DA release in contralateral structures. The most striking effect was seen withd-Ala₂,Met-Enk which enhanced tritiated DA release in the contralateral CN and SN. These results are discussed in the light of the involvement of several types of opiate receptors and of the polysynaptic pathways responsible for the distal changes in dopaminergic transmission.     

A role for striatal beta-adrenergic receptors in the regulation of dopamine release

The effect of (−) isoproterenol on the spontaneous release of [³H]dopamine ([³H]DA) continuously formed from [³H]tyrosine has been studied both in vitro using rat striatal slices and in vivo in halothane-anaesthetized cats implanted with push-pull cannulae in each caudate nucleus and each substantia nigra.In vitro, (−) isoproterenol (10⁻⁶M) stimulated [³H]DA release. This effect was stereospecific since (+) isoproterenol (10⁻⁶M) was without effect. (±) Propranolol (10⁻⁶M), which did not block the acetylcholine (10⁻⁵M)-induced release of [³H]DA, completely prevented the stimulatory effect of (−) isoproterenol on [³H]DA release. (±) Practolol (10⁻⁵M) significantly reduced the (−) isoproterenol-induced release of [³H]DA whereas phentolamine (10⁻⁵M) was uneffective. These results suggest that (−) isoproterenol acts on [³H]DA release through β₁-receptors. Since the adrenergic agonist still stimulated [³H]DA release in the presence of tetrodotoxin, the β-receptors involved may be located on DA terminals.Confirming in vitro results, (−) isoproterenol (10⁻⁶M) enhanced the local release of [³H]DA when applied into one caudate nucleus. This effect was biphasic and prevented by (±) propranolol (10⁻⁶M). The β-antagonist by itself slightly reduced [³H]DA spontaneous release suggesting that noradrenergic neurons which may innervate the caudate nucleus could exert a tonic facilitatory influence on DA release. No change in [³H]DA release occured in other structures studied during the application of (−) isoproterenol into one caudate nucleus.These various results indicate that striatal β-adrenergic receptors play a role in the regulation of DA release from DA nerve terminals and suggest an interaction between noradrenergic and dopaminergic neurones in the caudate nucleus.     

Triiodothyronine Regulates Insulin-Like Growth Factor-I Binding to Cultured Rat Pituitary Cells*

Triiodothyronine (T₃) stimulates the synthesis of growth hormone and enhances the growth of neoplastic rat pituitary somatomam-motrophs (GH cells) in culture. Moreover, T₃ has been shown to stimulate the production and secretion of an autocrine growth factor by these cells. We have previously demonstrated the presence of specific receptors for insulin-like growth factors (IGF) on GH cells. Since GH₃ cells contain mRNA encoding IGF-I, it has been suggested that IGF-I might act in an autocrine fashion in these cells. Therefore, it was of interest to learn how T₃ affects IGF-I binding to GH₃ cells. T₃ increased [¹²⁵I]IGF-I binding in a time - and dose-dependent manner. After 48 h of exposure to T₃, an increase in IGF-I binding was seen with 10^?11M T₃, maximizing with 10^?8M T₃. When cells were exposed to 10^?8 T₃, [¹²⁵I]IGF-I binding reached a maximum of 218 ± 20.8% of control (±SEM, P < 0.002) after 72 h of incubation. Scatchard analysis indicated that T₃ did not alter the K_d of IGF-I for its receptor, but that the total receptor number was increased. Dexamethasone (10^?7M) inhibited the T₃-induced increase in IGF-I binding, but glucocorticoid alone did not substantially alter receptor number. No significant change in insulin or IGF-II binding was seen after hormone treatment. 10^?8 M T₃ or IGF-I increased the growth of the GH₃ cells by ≥30%. Our data indicate that T₃ upregulates IGF-I binding in GH₃ cells without altering insulin binding and thereby provides a means for enhancing potential autocrine regulation in this cell line.     

Dopamine and 7-OH-DPAT may act on D3 receptors to inhibit tuberoinfundibular dopaminergic neurons

Whether the tuberoinfundibular dopaminergic (TIDA) neurons resided in the dorsomedial arcuate nucleus (dmARN) can respond to dopamine and a dopamine D₃ receptor agonist, 7-hydroxydipropylaminotetralin (7-OH-DPAT), was the focus of this study. In studies using extracellular single-unit recording of dmARN neurons in brain slices obtained from ovariectomized rats, dopamine and 7-OH-DPAT inhibited 60.1% (n = 141) and 80.9% (n = 47) of recorded dmARN neurons, respectively. Other dopamine D₁ or D₂ receptor agonists were not as effective. Intracerebroventricular injection of 7-OH-DPAT (10⁻⁹ mol/3 μl) in ovariectomized, estrogen-primed rats significantly lowered the TIDA neuronal activity as determined by 3,4-dihydroxyphenylacetic acid (DOPAC) levels in the median eminence. Co-administration of a putative D₃ receptor antagonist, U-99194A, could prevent the effect of 7-OH-DPAT. Unilateral microinjection of 7-OH-DPAT or dopamine itself (10⁻¹¹–10⁻⁹ mol/0.2 μl) into the right dmARN exhibited the same inhibitory effect on TIDA neurons. In all, dopamine may act on D₃ receptors to exhibit an inhibitory effect on its own release from the TIDA neurons.     

Regulation of α-melanocyte-stimulating hormone release from superfused slices of rat hypothalamus by serotonin and the interaction of serotonin with the dopaminergic system inhibiting peptide release

Release of α-melanocyte-stimulating hormone (α-MSH) from frontal slices of rat hypothalamus superfused with oxygenated artificial cerebrospinal fluid (ACSF) was quantified by radioimmunoassay. Control depolarisations with 50 mM KCl-containing ACSFm produced significant increases in α-MSH release which were partially blocked by 10⁻⁶ M cinnaserin, a serotonin (5-HT) receptor antagonist. Superfusion of the tissues with varying concentrations of 5-HT *10⁻⁷ M to 10⁻⁴ M) resulted in an inverted U-shaped dose-response curve, maximum α-MSH release being obtained with 10⁻⁶ M 5-HT. Addition of 10⁻⁶ M cinanserin shifted the 5-HT dose-response curve to the right whilst the presence of 10⁻⁸ M flupenthixol, a dopamine receptor antagonist, resulted in a sigmoidal 5-HT dose-response curve. Superfusion with ACSF containing either 10⁻⁷ M fluoxetine, a 5-HT re-uptake inhibitor, or 10⁻⁷ M p-chloroamphetamine, an agent releasing 5-HT, induced significant increases in α-MSH release which were abolished in the presence of 10⁻⁶ M cinanserin. These data demonstrate the presence of an endogenous 5-HT system that exerts a biphasic effect on α-MSH release. A stimulatory effect caused by lower 5-HT concentrations appears to be a direct action whilst an inhibitory effect at higher concentrations is mediated through an inhibitory endogenous dopaminergic system. A significant proportion of K⁺-stimulated peptide release is 5-HT-mediated.     

α-Adrenergic receptor-mediated depolarization of rat neocortical astrocytes in primary culture

The membrane potentials of rat neocortical astrocytes growing in primary cultures (mean resting potential; −79 mV at [K⁺]_o = 4.5 nM) were depolarized by up to 30 mV by 10⁻⁵ M norepinephrine added to the medium, or up to 11 mV by norepinephrine or phenylephrine applied by ionophoresis. This depolarization could be inhibited by the α-adrenergic receptor antagonist phentolamine (10⁻⁵ M) but not by the β-adrenergic antagonist propranolol (10⁻⁵ M). These results suggest that the norepinephrine-evoked depolarizations seen in these cells may be mediated through an α-adrenergic receptor.     

Evidence for a dopaminergic innervation of the cat lateral habenula: its role in controlling serotonin transmission in the Basal ganglia

The presence of a dopaminergic innervation of the cat lateral habenula and its possible role in modulating serotonin transmission within the basal ganglia were investigated using both in vitro and in vivo approaches. A high density of [³H]spiroperidol binding sites with similar affinities for domperidone and apomorphine as those present in the cat striatum were found in the habenula. By means of the push-pull cannula technique, a substantial release of [³H]dopamine continuously formed from [³H]tyrosine was detected in the lateral habenula of halothane-anesthetized cats since the amount of [³H]catecholamines was enhanced in the presence of benztropine, an inhibitor of dopamine uptake into dopaminergic nerve terminals. Furthermore, in anesthetized animals with a push-pull cannula implanted in each caudate nucleus and substantia nigra habenular applications of dopamine (10⁻⁷ M) reduced nigral but not striatal release of [³H]serotonin continuously formed from [³H]tryptophan. This change was prevented either by the delivery of domperidone to the lateral habenula or by the blockade of GABAergic transmission (picrotoxin 10⁻⁵ M) in the dorsal raphe. These data support the involvement of habenula-raphe pathways in the regulation of serotonin transmission in the cat basal ganglia and indicate that dopaminergic inputs to the lateral habenula participate in such a control.     

Modulation of glycine-induced currents by zinc and other metal cations in neurons acutely dissociated from the dorsal motor nucleus of the vagus of the rat

Effects of Zn²⁺ and other polyvalent cations on glycine-induced currents in the freshly dissociated rat dorsal motor nucleus of the vagus neurons were investigated under voltage-clamp conditions by the use of the nystatin-perforated patch recording configuration. Glycine evoked a Cl⁻ current in a concentration-dependent manner with a half-maximum effective concentration (EC₅₀) of 3.3×10⁻⁵ M. Strychnine inhibited the 3×10⁻⁵ M glycine-induced current in a concentration-dependent manner with a half-maximal inhibitory concentration (IC₅₀) of 6.8×10⁻⁷ M. At low concentrations (3×10⁻⁸ M–3×10⁻⁶ M), Zn²⁺ potentiated the current elicited by 3×10⁻⁵ M glycine. On the other hand, at concentrations higher than 10⁻⁵ M, Zn²⁺ inhibited the glycine response. The biphasic action of Zn²⁺ was mimicked by Ni²⁺, but La³⁺ and Co²⁺ had only potentiating effect. Zn²⁺ shifted the concentration–response curve for the glycine-induced current without changing the maximum response, and the EC₅₀ values for the glycine response in the absence and presence of 10⁻⁶ M and 10⁻⁴ M Zn²⁺ were 3.3×10⁻⁵ M, 1.3×10⁻⁵ M and 1.3×10⁻⁴ M, respectively. These results suggest that the biphasic modulation of glycine response by Zn²⁺ results from changes in apparent glycine affinity.     

Pituitary adenylate cyclase-activating polypeptide (PACAP) stimulates melatonin synthesis from rat pineal gland

The effect of pituitary adenylate cyclase polypeptide (PACAP) on rat pineal was examined. PACAP stimulated melatonin release from cultured dissociated pinealocytes with a 10⁴ higher potency than isoproterenol (EC₅₀ were 30 pM and 250 nM, respectively). The 10⁻⁹ M PACAP stimulation was not inhibited by 5 × 10⁻⁶ M VIP antagonist whereas that of 10⁻⁹ M VIP was reduced by 54%. Kinetic analysis of melatonin release indicated that PACAP acts postsynaptically via receptor activation.     

Effects of Ca and calmodulin on cyclic nucleotide metabolism in neurosecretosomes isolated from ox neurohypophyses

In neurosecretosomes, isolated from ox neurohypophyses, both guanylate and adenylate cyclase activity was shown to be predominantly membrane-bound. Membrane-bound adenylate cyclase was inhibited by increasing the ionized calcium concentration from 10⁻⁷ M to 10⁻⁵ M, but was stimulated by calmodulin in the presence of 10⁻⁷ M and 10⁻⁵ M ionized calcium. In contrast, neither calcium ions nor calmodulin affected the activity of membrane-bound guanylate cyclase. Soluble cyclic AMP and cyclic GMP phosphodiesterase activities increased with increasing ionized calcium concentration (10⁻⁷ M to 10⁻³ M). At 10⁻⁷ M ionized calcium concentration, both soluble phosphodiesterase activities were stimulated by calmodulin. Both the membrane-bound phosphodiesterase activities were inhibited by a high ionized calcium concentration (10⁻³ M) and not affected by calmodulin.     

Developmental studies of the uptake of choline, gaba and dopamine by crude synaptosomal preparations after in vivo or in vitro lead treatment

The kinetics of sodium dependent, high affinity uptake of choline and dopammine by striatal synaptosomal preparations and of GABA (gamma-aminobutyric acid) by cortical synaptosomal preparations have been examined during the development of Long-Evans control and lead-treated rats. Choline uptake was very low until 12 days postnatally, then the V_max increased and approached adult values of 29 pmol/mg prot./min within a week. GABA uptake was somewhat elevated at birth and only after three weeks did it decrease to the adult value of 0.7 nmol/mg prot./min. Dopamine uptake was low at birth, developed linearly with age and by 30 days postnatally approached the adult value of 68 pmoles/mg prot./min. The high affinity uptake constants (choline, 0.66 μM; GABA, 4.4 μM; and dopamine, 0.31 μM) did not change markedly during development. Similar studies were conducted with rats treated at the highest lead dosage which did not result in weight loss (100 μg lead as lead acetate/g body weight/day via intubation). Blood and brain lead determinations confirmed a substantial lead exposure. Such chronic exposure did not markedly affect the amount or developmental pattern of uptake of the putative neurotransmitters.The effect of 2.5 × 10⁻⁵ M lead acetate in vitro on the kinetics of high affinity uptake of these compounds into preparations from 20–26-day-old rats was investigated. When uptake was assayed in the absence of calcium, lead caused a 20% increase in the V_max for dopamine. This stimulation was reduced if samples were assayed in the presence of 1 mM CaCl₂. The K_m for high affinity uptake of these neurotransmitter-related compounds was not affected by lead.In other studies, crude synaptosomal preparations were preloaded with neurotransmitter by preincubation with radioactive choline, GABA, or dopamine. Release of radioactive neurotransmitter, either spontaneous or in response to potassium depolarization, was quantitated and correlated with the presence or absence of 2.5 × 10⁻⁵ M lead and/or 10⁻³ M calcium ions. Lead slightly inhibited calcium dependent spontaneous release of dopamine. Lead also appeared to partially substitute for calcium in the potassium depolarized release of dopamine and GABA, although subtraction of the spontaneous (potassium independent) component reduced the magnitude of the lead effect.     

Amphetamine-induced circling behavior in rats: Effects of unilateral microinjections of GABA and GABA-related drugs into substantia nigra

In these experiments, the GABA control upon dopaminergic nigrostriatal neurons has been investigated using circling behavior in the rat. Chronically cannulated rats were givend-amphetamine (2 mg/kg i.p.) 45 min before unilateral microinjection (0.2 μl/2 min) into the substantia nigra (SN) of GABA, muscimol, chlordiazepoxide (CDP) or bicuculline. Circling behavior was continuously recorded for 165 min using an automated rotometer. (1) Non-microinjected control rats exhibit a ‘spontaneous’ circling behavior after amphetamine. (2) When applied to the SN contralateral to the preferential side of the ‘spontaneous’ rotations, saline enhances contraversive circling; GABA (5 × 10⁻⁵M), CDP (5 × 10⁻⁵M) and muscimol (5 × 10⁻⁷M, 5 × 10⁻⁸M) counteract this effect and induce (except CDP) light ipsiversive rotations; GABA (10⁻²M) and muscimol (5 × 10⁻⁵M) further enhance contraversive turning. (3) When applied to the SN ipsilateral to the preferential side of the ‘spontaneous’ rotations, saline has no marked effect on the ipsiversive circling behavior but induces weak contraversive turning; GABA (5 × 10⁻⁵M), CDP (5 × 10⁻⁵M) and muscimol (5 × 10⁻⁸M) enhance the ipsiversive rotations; GABA (10⁻²M) and muscimol (5×10⁻⁵M) transiently decrease the ipsiversive circling; bicuculline (5 × 10⁻⁵M) induces a vigorous contraversive turning associated with a transient inhibition of the ipsiversive rotations. These results suggest that the activity of the nigral neurons is presumably stimulated by the microinjection itself, by bicuculline and, to a lesser extent, by high concentrations of GABA and muscimol, and inhibited by low concentrations of GABA, muscimol and CDP. These findings could further support the hypotheses of a GABAergic inhibitory control upon DA nigrostriatal pathways and of a GABA-like activity of CDP.     

Dopamine released from dendrites in the substantia nigra controls the nigral and striatal release of serotonin

Using push-pull cannulae, the release of endogenously synthesized [³H]serotonin was estimated in both substantia nigra and caudate nuclei of ‘encéphale isolé’ cats. The unilateral nigral application of dopamine (10⁻⁷ M) reduced [³H]serotonin release in ipsilateral structures whereas α-methylparatyrosine (10⁻⁴ M) induced opposite effects. Both treatments decreased [³H]serotonin release in the contralateral caudate nucleus but not in the contralateral substantia nigra. As a working hypothesis it is suggested that the effects of observed are related to changes in the activity of nigroraphe neurons regulated by dopamine released from dendrites of the nigrostriatal dopaminergic neurons. However it cannot vet be excluded that the local changes in [³H]serotonin release induced by the nigral application of dopamine or α-methylparatyrosine result from presynaptic modulation.     

GABA-ergic control of α-Melanocyte-Stimulating Hormone (α-MSH) release by frog neurointermediate lobe in vitro

Measurement of glutamate decarboxylase (GAD) activity in the intermediate lobe of the frog pituitary and brain showed that neurointermediate lobe extracts represented 12% of the GAD activity detected in the whole brain. No significant activity was measured in distal lobe extracts. Immunocytochemical studies revealed GAD-containing fibers among the parenchyma! cells of the pars intermedia. The localization of GAD-like material in the intermediate lobe of the frog pituitary suggested a possible role of γ-aminobutyric acid (GABA) in the regulation of melanotropic cell secretion. Administration of GAB A (10⁻⁶ to 10⁻⁴ M), to perifused neurointermediate lobes caused a brief stimulation of alpha-melanocyte stimulating hormone (α-MSH) release followed by an inhibition. Picrotoxin (10⁻⁴ M), a Cl⁻ channel blocker, abolished only the stimulatory effect of GAB A (10⁻⁴ M), whereas bicuculline (10⁻⁴ M), a specific antagonist of GABA_A receptors, totally inhibited the effects of GABA (both stimulatory and inhibitory phases). Bicuculline induced by itself a slight stimulation of α-MSH release, suggesting that GABA-ergic nerve fibers present in the intermediate lobe are functionally active in vitro. The GABA_A agonist muscimol (10⁻⁷ to 10⁻⁴ M) mimicked the biphasic effect of GABA on α-MSH release. Administration of baclofen, a specific GABA_BB agonist (10⁻⁷ to 10⁻⁴ M) induced a dose-dependent inhibition of α-MSH secretion. In contrast to GABA or muscimol, baclofen did not cause any stimulatory effect whatever the dose. Taken together these result suggested that GABA_A and GABa_b receptors were present on frog melanotrophs. Since bicuculline totally inhibited GABA effects (stimulation and inhibition) on α-MSH release, it appears however that the effect of GABA is mainly achieved through activation of G AB A_A receptors.