ACTH-(1-24) and alpha-MSH antagonize feeding behavior stimulated by kappa opiate agonists

ACTH-(1-24) and alpha-MSH, intracerebroventricularly (ICV) injected at the doses of 4 and 10 micrograms/animal, respectively, markedly inhibited spontaneous feeding in adult Sprague-Dawley rats, the effect remaining significant for 6-9 hours. At these same doses, ACTH-(1-24) and alpha-MSH abolished the feeding-stimulatory effect of the kappa opiate receptor agonist pentazocine, intraperitoneally (IP) injected at the dose of 10 mg/kg. The same antagonism was obtained by ICV injection of ACTH-(1-24) into rats IP treated with other kappa opiate agonists, bremazocine and tifluadom, at the doses of 1 and 5 mg/kg, respectively. These data suggest that melanocortin peptides play an inhibitory role in the complex regulation of food intake, and further support and extend the hypothesis of a melanocortin-opioid homeostatic system, its two neuropeptide components usually having opposite, mutually-balancing effects.     

Influence of Cannabinoids on Electrically Evoked Dopamine Release and Cyclic AMP Generation in the Rat Striatum

Abstract: Using the endogenous cannabinoid receptor agonist anandamide, the synthetic agonist CP 55940 {[1α,2β( R )5α]-(−)-5-(1,1-dimethylheptyl)-2-[5-hydroxy-2-(3-hydroxypropyl)cyclohexyl]phenol}, and the specific antagonist SR 141716 [ N -(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1 H -pyrazole-3-carboxamide hydrochloride], second messenger activation of the central cannabinoid receptor (CB1) was examined in rat striatal and cortical slices. The effects of these cannabinoid ligands on electrically evoked dopamine (DA) release from [³H]dopamine-prelabelled striatal slices were also investigated. CP 55940 (1 µ M ) and anandamide (10 µ M ) caused significant reductions in forskolin-stimulated cyclic AMP accumulation in rat striatal slices, which were reversed in the presence of SR 141716 (1 µ M ). CP 55940 (1 µ M ) had no effect on either KCl- or neurotransmitter-stimulated ³H-inositol phosphate accumulation in rat cortical slices. CP 55940 and anandamide caused significant reductions in the release of dopamine after electrical stimulation of [³H]dopamine-prelabelled striatal slices, which were antagonised by SR 141716. SR 141716 alone had no effect on electrically evoked dopamine release from rat striatal slices. These data indicate that the CB1 receptors in rat striatum are negatively linked to adenylyl cyclase and dopamine release. That the CB1 receptor may influence dopamine release in the striatum suggests that cannabinoids play a modulatory role in dopaminergic neuronal pathways.     

Structure-activity studies with ACTH/alpha-MSH fragments on corticosteroid secretion of isolated zona glomerulosa and fasciculata cells

The steroidogenic action of ACTH/alpha-MSH fragments was studied on isolated zona glomerulosa and zona fasciculata cells dispersed by collagenase. ACTH-(4-7), ACTH-(6-10), ACTH-(4-10) and ACTH-(11-13) stimulated corticosterone production of the zona fasciculata and aldosterone production of the zona glomerulosa cells. ACTH-(7-10) was ineffective. ACTH-(4-7) appeared to be the most potent peptide of the tested fragments. None of the fragments affected the steroidogenic action of ACTH-(1-39). It is suggested that similar to the melanotropic effect of alpha-MSH two 'message' sequences for adrenocortical stimulation exist in the alpha-MSH part of the ACTH molecule.     

Regulation of pro-opiomelanocortin synthesis by dopamine and cAMP in the amphibian pituitary intermediate lobe

The modulation of pro-opiomelanocortin (POMC) synthesis in Xenopus laevis pituitary intermediate lobe (IL) during background adaptation and the role of dopamine and cAMP in mediating this effect were examined. Neurointermediate lobes (NILs) were pulselabeled in vitro with [3H]arginine and analyzed for POMC synthesis by acid-urea gel electrophoresis. After black background adaptation of the animal (7 days), POMC synthesis increased 5-6-fold, while after white background adaptation (7 days), POMC synthesis decreased by 76%. Dopamine (50 microM) suppressed POMC synthesis in NILs in culture. In the absence of dopamine, POMC synthesis was stimulated. Several experiments were conducted to determine the category of dopamine receptor in the X. laevis IL. A D-2 dopamine receptor agonist inhibited immunoreactive alpha-MSH release from the NIL in a D-2 antagonist-reversible manner. A D-1 receptor agonist or antagonist did not alter the release of immunoreactive alpha-MSH from the NIL. Dopamine (10 microM) inhibited forskolin-stimulated cAMP accumulation. In addition, dopamine inhibition of POMC synthesis in cultured ILs was reversed by 8-Br-cAMP. These studies suggest that white background adaptation results in stimulation of the X. laevis D-2 receptor, which reduces cAMP production and POMC synthesis. Conversely, during black background adaptation the IL D-2 receptor is not stimulated, leading to increased cAMP production and POMC synthesis.     

The benzodiazepine agonist clonazepam potentiates the effects of gamma-aminobutyric acid on alpha-MSH release from neurointermediate lobes in vitro

The action of the central-type benzodiazepine-receptor agonist clonazepam on alpha-MSH release has been studied in vitro using perifused frog neurointermediate lobes. High concentrations of clonazepam (3.16 X 10(-5) and 10(-4) M) caused an inhibition of alpha-MSH release and this effect was reversed by the central-type benzodiazepine-receptor antagonist Ro 15-1788. High doses of GABA (10(-5) and 10(-4) M) induced a biphasic effect on pars intermedia cells: a brief stimulation followed by a sustained inhibition of alpha-MSH secretion. Administration of clonazepam (10(-5) M) in the presence of various concentrations of GABA (10(-6) to 10(-4) M) led to a potentiation of both stimulatory and inhibitory phases of alpha-MSH secretion induced by GABA. Ro 15-1788 completely abolished the potentiating effect of clonazepam. Thus our results indicate that endogenous benzodiazepine receptors may modulate the effects of GABA on alpha-MSH secretion.     

Effects of L-dopa and L-tyrosine on release of free and conjugated dopamine, homovanillic acid and dihydroxyphenylacetic acid from slices of rat striatum

Conjugation (presumably with sulfate) is a demonstrable metabolic pathway for 3, 4-dihydroxyphenylethylamine (dopamine, DA) in brain. Studies were done to determine whether conjugation becomes of increased significance in the presence of precursors of DA. The effects of 3, 4-dihydroxyphenylalanine (L-DOPA) and L-tyrosine on the efflux of free and conjugated DA, 3, 4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid from slices from striatum in rats were studied under quiescent conditions and during release evoked by 40 mM K+ or by 5 X 10(-5) M phenylethylamine (PEA). Conjugated DA was present in the basal efflux from striatal slices and the amounts present were increased during evoked release. More conjugated DA was present in superfusate during K+-evoked release than during PEA-evoked release. L-Tyrosine (5 X 10(-4) M or 5 X 10(-5) M) had little effect on the efflux of conjugated DA, but decreased the amounts of free DA released by PEA, and attenuated the increase in DOPAC that occurred during K+-evoked release of transmitter. L-DOPA (5 X 10(-5) M) increased the formation of conjugated DA, but to a lesser extent than that of free DA or of DOPAC. Thus even after the addition of precursors, conjugation remains a minor metabolic pathway for DA relative to O-methylation or oxidative deamination. The data also suggest that conjugation of DA occurs chiefly outside of the dopaminergic neurons in striatum.     

Effect of melanostatin and thyroliberin on the biosynthesis and release of dopamine by rat brain striatal P2 fractions

The effect of melanostatin (MIF) and thyroliberin (TRH) on dopamine biosynthesis and release has been studied in striatal P₂ fractions from rat brain. Dopamine synthesis was not affected by either MIF (10^?4M) or TRH (10^?4M) in the presence of 2 mM EDTA. In contrast, tyramine (10^?4M) and (+)-amphetamine (10^?4M) both stimulated dopamine biosynthesis, in a manner inversely proportional to the rate of dopamine biosynthesis, in the absence of drugs. MIF (10^?3M) or TRH (10^?3M) stimulated the release of dopamine in the presence of 1.25 mM calcium chloride but not in the presence of 2 mM EDTA. In contrast (+)-amphetamine 10^?4M) stimulated release of dopamine under both conditions. These results are discussed in relation to the hypothesis that MIF and TRH stimulate the release of dopamine from vesicular stores by a calcium dependent mechanism, whereas amphetamine stimulates release of dopamine from both vesicular stores and a cytoplasmic pool which regulates tyrosine hydroxylase activity.     

Involvement of KCNQ2 subunits in [3H]dopamine release triggered by depolarization and pre-synaptic muscarinic receptor activation from rat striatal synaptosomes

KCNQ2 and KCNQ3 subunits encode for the muscarinic-regulated current (I(KM)), a sub-threshold voltage-dependent K+ current regulating neuronal excitability. In this study, we have investigated the involvement of I(KM) in dopamine (DA) release from rat striatal synaptosomes evoked by elevated extracellular K+ concentrations ([K+]e) and by muscarinic receptor activation. [3H]dopamine ([3H]DA) release triggered by 9 mmol/L [K+]e was inhibited by the I(KM) activator retigabine (0.01-30 micromol/L; Emax = 54.80 +/- 3.85%; IC50 = 0.50 +/- 0.36 micromol/L). The I(KM) blockers tetraethylammonium (0.1-3 mmol/L) and XE-991 (0.1-30 micromol/L) enhanced K+-evoked [3H]DA release and prevented retigabine-induced inhibition of depolarization-evoked [3H]DA release. Retigabine-induced inhibition of K+-evoked [3H]DA release was also abolished by synaptosomal entrapment of blocking anti-KCNQ2 polyclonal antibodies, an effect prevented by antibody pre-absorption with the KCNQ2 immunizing peptide. Furthermore, the cholinergic agonist oxotremorine (OXO) (1-300 micromol/L) potentiated 9 mmol/L [K+]e-evoked [3H]DA release (Emax = 155 +/- 9.50%; EC50 = 25 +/- 1.80 micromol/L). OXO (100 micromol/L)-induced [3H]DA release enhancement was competitively inhibited by pirenzepine (1-10 nmol/L) and abolished by the M3-preferring antagonist 4-diphenylacetoxy N-methylpiperidine methiodide (1 micromol/L), but was unaffected by the M1-selective antagonist MT-7 (10-100 nmol/L) or by Pertussis toxin (1.5-3 microg/mL), which uncouples M2- and M4-mediated responses. Finally, OXO-induced potentiation of depolarization-induced [3H]DA release was not additive to that produced by XE-991 (10 micromol/L), was unaffected by retigabine (10 micromol/L), and was abolished by synaptosomal entrapment of anti-KCNQ2 antibodies. Collectively, these findings indicate that, in rat striatal nerve endings, I(KM) channels containing KCNQ2 subunits regulate depolarization-induced DA release and that I(KM) suppression is involved in the reinforcement of depolarization-induced DA release triggered by the activation of pre-synaptic muscarinic heteroreceptors.     

Dopamine Inhibition of the Release of Endogenous Acetylcholine from Corpus Striatum and Cerebral Cortex in Tissue Slices and Synaptosomes: A Presynaptic Response?

Abstract: The effect of dopamine on the release of endogenous acetylcholine from striatal slices and synaptosomes and from cerebral cortex synaptosomes was studied. K⁺ (56 m M ) and veratrine (75 μM ) increased the release of acetylcholine from striatal slices by 3.7 and 3.3 times the resting release, respectively. The effect of veratrine was completely abolished by tetrodotoxin (1 μM ). Dopamine (10⁻⁶ to 10⁻³ M ) reduced the K⁺-evoked release of acetylcholine from striatal slices in a dose-dependent manner. The resting release of acetylcholine was also significantly reduced by dopamine. Apomorphine (20 μM ) significantly reduced the K⁺-evoked release of acetylcholine, and both this effect and the inhibition due to dopamine (1 m M ) were significantly antagonised by chlorpromazine (20 μM ). Dopamine had a similar effect on the release of acetylcholine from striatal synaptosome beds; the resting release was depressed 32% by the presence of dopamine (1 m M ). A greater effect of dopamine was seen on the release of acetylcholine from cerebral cortex synaptosome beds, the resting release being reduced by 54% and the K⁺-evoked release by 29%. These results are discussed in terms of the possible role of presynaptic dopamine receptors in controlling the release of acetylcholine and the magnitude of their contribution compared with that of the postsynaptic dopamine receptor.     

Persisting subsensitivity of the striatal dopamine system after fetal exposure to beta-endorphin

Fetal exposure of rats to beta-endorphin during the third trimester, either alone or with alpha-MSH, resulted in mild developmental delay and significant decreases in striatal dopamine receptor density (subsensitivity) persisting through maturity. The apparent paradoxical down-regulation of dopamine receptors in the presence of beta-endorphin was consistent with fetal exposure to dopamine receptor antagonists and synthesis inhibitors. These findings suggest biophysical properties of receptors which are unique to fetal development including loss of plasticity after exposure to antagonists. Permanent, down-regulation of the striatal dopamine system may be one mechanism underlying delayed development after fetal exposure to beta-endorphin which may accompany hypoxia. Even though there were no statistically significant differences between males and females in density of the dopamine receptor, the behavioral profile after peptide treatment was sexually demorphic. Behaviorally, female rats appeared sensitized to perinatal alpha-MSH and males to alpha-endorphin.     

Subsecond regulation of striatal dopamine release by pre-synaptic KATP channels

ATP-sensitive K(+) (K(ATP)) channels are composed of pore-forming subunits, typically Kir6.2 in neurons, and regulatory sulfonylurea receptor subunits. In dorsal striatum, activity-dependent H(2)O(2) produced from glutamate receptor activation inhibits dopamine release via K(ATP) channels. Sources of modulatory H(2)O(2) include striatal medium spiny neurons, but not dopaminergic axons. Using fast-scan cyclic voltammetry in guinea-pig striatal slices and immunohistochemistry, we determined the time window for H(2)O(2)/K(ATP)-channel-mediated inhibition and assessed whether modulatory K(ATP) channels are on dopaminergic axons. Comparison of paired-pulse suppression of dopamine release in the absence and presence of glibenclamide, a K(ATP)-channel blocker, or mercaptosuccinate, a glutathione peroxidase inhibitor that enhances endogenous H(2)O(2) levels, revealed a time window for inhibition of 500-1000 ms after stimulation. Immunohistochemistry demonstrated localization of Kir6.2 K(ATP)-channel subunits on dopaminergic axons. Consistent with the presence of functional K(ATP) channels on dopaminergic axons, K(ATP)-channel openers, diazoxide and cromakalim, suppressed single-pulse evoked dopamine release. Although cholinergic interneurons that tonically regulate dopamine release also express K(ATP) channels, diazoxide did not induce the enhanced frequency responsiveness of dopamine release seen with nicotinic-receptor blockade. Together, these studies reveal subsecond regulation of striatal dopamine release by endogenous H(2)O(2) acting at K(ATP) channels on dopaminergic axons, including a role in paired-pulse suppression.     

Adrenocorticotropin/α-Melanocyte-Stimulating Hormone (ACTH/MSH)-Like Peptides Modulate Adenylate Cyclase Activity in Rat Brain Slices: Evidence for an ACTH/MSH Receptor-Coupled Mechanism

Abstract: The regulation of adenylate cyclase activity by adrenocorticotropin/α-melanocyte–stimulating hormone (ACTH/MSH)-like peptides was investigated in rat brain slices using a superfusion method. Adenylate cyclase activity was concentration-dependently increased by ACTH-(1–24), α-MSH (EC₅₀ values 16 and 6 nM, respectively), and [Nle⁴,D-Phe⁷]α-MSH (EC₅₀ value 1.6 nM), in the presence of forskolin (1 μM, optimal concentration). 1-9-Dideoxy-forskolin did not augment the response of adenylate cyclase to ACTH-(1–24). Various peptide fragments were tested for their ability to enhance [³H]cyclic AMP production. [Nle⁴,D-Phe⁷]α-MSH increased [³H]cyclic AMP formation with a maximal effect of 30% and was more potent than ACTH-(1–24), ACTH-(1–16)-NH₂, α-MSH, ACTH-(1–13)-NH₂, [MetO⁴]α-MSH, [MetO₂⁴,D-Lys⁸,Phe⁹]ACTH-(4–9), ACTH-(7–16)-NH₂, ACTH-(1–10), and ACTH-(11–24), in order of potency. This structure–activity relationship resembles that found for the previously described peptide-induced display of excessive grooming. ACTH-(1–24) stimulated adenylate cyclase activity in both striatal (maximal effect, ?20%) and septal slices (maximal effect, ?40%), but not in hippocampal or cortical slices. Lesioning of the dopaminergic projections to the striatum did not result in a diminished effect of [Nle⁴,D-Phe⁷]α-MSH on [³H]cyclic AMP accumulation, which indicates that the ACTH/MSH receptor–stimulated adenylate cyclase is not located on striatal dopaminergic terminals. ACTH-(1–24) did not affect the dopamine D₁ or D₂ receptor–mediated modulation of adenylate cyclase activity. Based on the present data, we suggest that the binding of endogenous ACTH or α-MSH to a putative ACTH/MSH receptor in certain brain regions leads to the activation of a signal transduction pathway using cyclic AMP as a second messenger.     

Evidence for a functional coupling between dopamine reuptake and tyrosine hydroxylation in striatal nerve terminals

In rat striatal synaptosomes incubated with [¹⁴C]tyrosine, the evolution of ¹⁴CO₂, taken as a measure of dopamine synthesis, was inhibited by exogenous dopamine and by the dopaminergic receptor agonist ADTN. The inhibition was not counteracted by dopaminergic receptor antagonists (haloperidol, sulpiride, pimozide or domperidone). Instead, it was prevented by dopamine uptake blockers, suggesting that dopamine and ADTN (a substrate of the dopamine carrier) acted once inside the nerve endings and not through activation of autoreceptors on their external membrane. The dopamine uptake inhibitors nomifensine, benztropine and cocaine increased ¹⁴CO₂ evolution from incubated striatal synaptosomes. Depolarization with KCl also increased dopamine synthesis and this action was potentiated when the reuptake of the released catecholamine was prevented by carrier blockers. The rate of dopamine synthesis was lowered when synaptosomal dopamine was raised upon incubation with monoamine oxidase inhibitors or with l-DOPA. The inhibition was counteracted by dopamine reuptake blockers. The data suggest that dopamine synthesis in striatal nerve endings is under the inhibitory control of the transmitter recaptured following release.     

Exocytotic release of dopamine in ventral tegmental area slices from C57BL/6 and dopamine transporter knockout mice

The present study used voltammetry to ascertain whether electrically stimulated somatodendritic dopamine release in ventral tegmental area slices from C57BL/6 and dopamine transporter knockout mice was due to exocytosis or dopamine transporter reversal, as has been debated. The maximal concentration of electrically evoked dopamine release was similar between ventral tegmental area slices from dopamine transporter knockout and C57BL/6 mice. Dopamine transporter blockade (10 μM nomifensine) in slices from C57BL/6 mice inhibited dopamine uptake but did not alter peak evoked dopamine release. In addition, dopamine release and uptake kinetics in ventral tegmental area slices from dopamine transporter knockout mice were unaltered by the norepinephrine transporter inhibitor, desipramine (10 μM), or the serotonin transporter inhibitor, fluoxetine (10 μM). Furthermore, maximal dopamine release in ventral tegmental area slices from both C57BL/6 and dopamine transporter knockout mice was significantly decreased in response to Na⁺ channel blockade by 1 μM tetrototoxin, removal of Ca²⁺ from the perfusion media and neuronal vesicular monoamine transporter inhibition by RO-04-1284 (10 μM) or tetrabenazine (10 and 100 μM). Finally, the glutamate receptor antagonists AP-5 (50 and 100 μM) and CNQX (20 and 50 μM) had no effect on peak somatodendritic dopamine release in C57BL/6 mice. Overall, these data suggest that similar mechanisms, consistent with exocytosis, govern electrically evoked dopamine release in ventral tegmental area slices from C57BL/6 and dopamine transporter knockout mice.     

Interaction between alpha-MSH and acetylcholinergic system upon striatal cAMP and IP(3) levels

The interaction between the neuropeptide alpha-MSH and the acetylcholinergic system as reflected by changes in cAMP and inositol 1-3-5 triphosphate(IP(3))production was investigated in an in vitro model of striatal slices. The possible involvement of D(1) receptors in cholinergic and alpha-MSH- stimulated cAMP and IP(3) production in slices of rat striatum was also examined, because it has been demonstrated that acetylcholinergic drugs induce endogenous dopamine release in the striatum. alpha-MSH, pilocarpine(PL) and the selective muscarinic M1 agonist McN-A-343 increased cAMP and IP(3) striatal levels, effects blocked by the D(1) antagonist SCH-23390, except for the effects of alpha-MSH on IP(3).The muscarinic M(2) antagonist gallamine (GL) brought about an increase in cAMP levels, an effect blocked by SCH-23390. The M(1) antagonist pirenzepine (Pz) induced a decrease both in cAMP and IP(3) content, and the nicotinic antagonist di-hydro-beta-eritroidine(DBE) only diminished cAMP production. When alpha-MSH and cholinergic agents were simultaneously added, cAMP and IP(3) levels were modified with respect to the values reached when these agents were added alone. An interaction between the acetylcholinergic system and alpha-MSH through M(1) and nicotinic receptors was also observed. These results suggest that the intracellular signaling pathways related to cAMP and IP(3) production gated by alpha-MSH and these cholinergic receptors are probably related. alpha-MSH striatum cAMP IP(3) muscarinic and nicotinic receptors an in vitro model.     

Amperozide, a putative anti-psychotic drug: uptake inhibition and release of dopamine in vitro in the rat brain

The effects of amperozide (a diphenylbutylpiperazinecarboxamide derivative) on the uptake and release of 3H-dopamine in vitro were investigated. Amperozide inhibited the amphetamine-stimulated release of dopamine from perfused rat striatal tissue in a dose-dependent manner. With 1 and 10 microM amperozide there was significant inhibition of the amphetamine-stimulated release of dopamine, to 44 and 36% of control. In contrast, 10 microM amperozide significantly strengthened the electrically stimulated release of dopamine from perfused striatal slices. Amperozide 1-10 microM had no significant effect on the potassium-stimulated release of dopamine. 10 microM amperozide also slightly increased the basal release of 3H-dopamine from perfused striatal tissue. These effects on various types of release are similar to those reported for uptake inhibitors (Bowyer et al, 1984). The uptake of dopamine in striatal tissue was inhibited by amperozide with IC50 values of 18 microM for uptake in chopped tissue and 1.0 microM for uptake in synaptosomes. Amperozide also inhibited the uptake of serotonin in synaptosomes from frontal cortex, IC50 = 0.32 microM and the uptake of noradrenaline in cortical synaptosomes, IC50 = 0.78 microM. In conclusion, amperozide shows uptake-inhibiting properties in both release and uptake studies done in vitro on the rat. In the in vivo studies, however, amperozide differs from dopamine uptake inhibitors.     

Dopaminergic regulation of glucose-induced insulin secretion through dopamine D2 receptors in the pancreatic islets in vitro

The stimulatory effect of dopamine through dopamine D2 receptor on glucose-induced insulin secretion was studied in the pancreatic islets in vitro. Dopamine significantly stimulated insulin secretion at a concentration of 10-8 M in the presence of high glucose (20 mM). The higher concentrations of dopamine (10(-7)-10(-4)) inhibited glucose-induced insulin secretion in the presence of both 4 mM and 20 mM glucose. Stimulatory and inhibitory effect of dopamine on glucose-induced insulin secretion was reverted by the addition of dopamine D2 receptor antagonists such as butaclamol and sulpiride. Norepinephrine (NE) at 10(-4) M concentration inhibited the dopamine uptake as well as its stimulatory effect at 10(-8) M concentration on glucose induced insulin secretion. Our results suggest that dopamine exerts a differential effect on glucose-induced insulin secretion through dopamine D2 receptor and it is essential for the regulation of glucose-induced insulin secretion by pancreatic islets.     

Both dopamine and the putative dopamine D3 receptor antagonist PNU-99194A induce a biphasic inhibition of phorbol ester-stimulated arachidonic acid release from CHO cells transfected with the dopamine D3 receptor

In Chinese hamster ovary (CHO) cells transfected with the cDNA for the dopamine D3 receptor, low concentrations of dopamine (IC50: 0.5 nM) counteracted the release of arachidonic acid (AA) induced by the protein kinase C activator TPA (maximal inhibition: 15% at 10 - 30 nM). The effect of dopamine -- which was antagonized by pretreatment with pertussis toxin (PTX) or by the dopamine receptor antagonist haloperidol -- was biphasic; thus, at increasing concentrations of dopamine (100 nM - 1 microM), AA levels approached baseline. The preferential dopamine D3 receptor ligand PNU-99194A displayed an effect similar to that of dopamine; thus, whereas low concentrations of PNU-99194A (IC50: 1.9 nM) reduced TPA-induced AA release (maximal inhibition: 15% at 30 - 100 nM), higher concentrations (> or =1 microM) were ineffective. When dopamine and PNU-99194A were administered together at concentrations yielding maximal inhibition of AA release, no additive effect was observed; moreover, a high concentration of dopamine counteracted the AA-reducing effect of a low concentration of PNU-99194A and vice versa. It is suggested that D3 receptors in transfected CHO cells may exert mainly an inhibitory, but also a stimulatory influence on TPA-induced AA release, and that PNU-99194A acts as an agonist in this system.     

Presynaptic Dopamine Autoreceptors Control Tyrosine Hydroxylase Activation in Depolarized Striatal Dopaminergic Terminals

The possible control of tyrosine hydroxylase (TH) activity by dopaminergic receptor-dependent mechanisms was investigated using rat striatal slices or synaptosomes incubated in the presence of various 3,4-dihydroxyphenylethylamine (dopamine or DA) agonists and antagonists. Under "normal" conditions (4.8 mM K+ in the incubating medium), the DA agonists apomorphine, 6,7-dihydroxy-N,N-dimethyl-2-aminotetralin (TL-99), 7-hydroxy-N,N-dipropyl-2-aminotetralin (7-OH-DPAT), Trans-(-)-4,4a,5,6,7,8,8a,9-octahydro-5-propyl-2H-pyrazolo-3,4- quinoline, and 3-(3-hydroxyphenyl)-N-n-propylpiperidine decreased TH activity in soluble extracts of incubated tissues. In the case of the catechol-containing drugs apomorphine and TL-99, this effect was partly due to a direct inhibition of the enzyme, but in all other cases it appeared to depend on the stimulation of presynaptic DA autoreceptors. No effect of DA antagonists was detected on TH activity under "normal" conditions. In contrast, when tissues were incubated in a K+ -enriched (60 mM) medium, (-)-sulpiride and other DA antagonists enhanced TH activation due to depolarization whereas DA agonists were ineffective. Because (-)-sulpiride also increased the enzyme activity in striatal slices exposed to drugs inducing release of DA, such as veratridine and d-amphetamine, it is concluded that the stimulating effect of the DA antagonist resulted in fact from the blockade of the negative control of TH normally triggered by endogenous DA acting on presynaptic autoreceptors. In contrast to TH activation due to K+ -induced depolarization, the activation evoked by tissue incubation with dibutyryl cyclic AMP was unaffected by the typical agonist 7-OH-DPAT or the antagonist (-)-sulpiride. This would suggest that TH control via presynaptic DA autoreceptors normally concerns possible modulations of the cyclic AMP-dependent phosphorylation of the enzyme.     

Functional antagonism of D-1 and D-2 dopaminergic mechanisms affecting striatal acetylcholine release

Rat striatal slices prelabelled with [3H]choline were superfused with dopamine D-1 and D-2 agonists and antagonists, separately and in combination, during measurement of [3H]acetylcholine (ACh) release. SKF38393 (D-1 agonist), 10(-7)-10(-4) M, and SCH23390 (D-1 antagonist), 10(-7)-10(-5) M, produced a dose-dependent increase in [3H]ACh release when given separately. The increased [3H]ACh release induced by either drug could not be attenuated by sufficient L-sulpiride to block D-2 receptors. Yet both SKF38393, 10(-6)-10(-5) M, and SCH23390, 10(-6)-10(-5) M, were able to partially or fully overcome the [3H]ACh release-depressant effect of cosuperfused LY171555 (D-2 agonist), 10(-6) M. This suggests that a functional antagonism regarding striatal ACh release exists between D-1 and D-2 dopaminergic receptor-mediated mechanisms, but that D-1 modulation of local ACh release does not occur at the level of the recognition site of the striatal D-2 receptor. Finally, although attenuation of the increased release of striatal [3H]ACh induced by 10(-5) M SCH23390 by SKF38393 was seen, it is possible that such functional antagonism is not mediated by exclusively D-1 dopaminergic means.