Differential Effects of D1 and D2 Dopamine Receptor Antagonists on Acute Amphetamine- or Methamphetamine-Induced Up-Regulation of zif/268 mRNA Expression in Rat Forebrain

Abstract: This study investigated the hypothesis that D₁ and D₂ dopamine receptors interact to regulate the expression of zif/268 mRNA in rat forebrain after an acute injection of amphetamine or methamphetamine. Forty-five minutes and 3 h after a single injection of amphetamine (4 mg/kg i.p.) or methamphetamine (4 mg/kg i.p.), the mRNA expression of zif/268 in dorsal striatum and sensorimotor cortex was increased, as revealed by quantitative in situ hybridization histochemistry. Induction was more intense in striatal patches at 45 min than at 3 h, when a more homogeneous pattern of zif/268 mRNA induction was observed. SCH 23390, a selective D₁ receptor antagonist, suppressed, and eticlopride, a D₂ receptor antagonist, elevated, constitutive zif/268 mRNA levels in the striatum, but neither antagonist had a significant effect on the constitutive expression of zif/268 in sensorimotor cortex. Pretreatment with SCH 23390 completely blocked the stimulant-induced zif/268 expression in striatum and partially blocked the stimulant-induced zif/268 expression in cortex. Pretreatment with eticlopride augmented zif/268 mRNA expression in patch and matrix compartments of dorsal and ventral striatum 45 min after amphetamine or methamphetamine injection. However, at 3 h, eticlopride completely blocked amphetamine- and methamphetamine-stimulated zif/268 mRNA in dorsomedial, but not dorsolateral, striatum. In addition, eticlopride partially blocked cortical zif/268 induction by both amphetamines. Both antagonists prevented stimulant-induced hyperlocomotion and stereotypies. These results demonstrate that D₁ and D₂ receptors in mesolimbic/mesostriatal pathways both regulate amphetamine-and methamphetamine-stimulated behaviors and zif/268 mRNA expression. Furthermore, the effect of D₂ receptor blockade on zif/268 expression was found to be contingent on the time interval investigated after psychostimulant administration.     

D1 Dopamine Receptor Binding and mRNA Levels Are Not Altered After Neonatal 6-Hydroxydopamine Treatment: Evidence Against Dopamine-Mediated Induction of D1 Dopamine Receptors During Postnatal Development

Abstract: The role of dopaminergic innervation on the postnatal developmental expression of D₁ dopamine receptors was investigated. Bilateral destruction of dopa-mine-containing neurons was achieved by treating rats intracisternally with 6-hydroxydopamine (6-OHDA) on postnatal day 3, and rats were killed on day 21. To ensure effective reduction of D₁ receptor activation by residual dopamine, a group of 6-OHDA-lesioned rats was given twice daily injections of the D₁ receptor antagonist SCH-23390, from day 4 to 20. D₁ dopamine receptor binding was assessed in the caudate—putamen, nucleus accumbens, and olfactory tubercle by quantitative autoradiographic analysis of [³H]SCH-23390 binding. In addition, the relative amount of D_1A receptor mRNA was assessed by in situ hybridization of a ³⁵S-labeled riboprobe. In the developing rats, neither the amount of [³H]SCH-23390 binding nor the amount of D_1A receptor mRNA was altered by 6-OHDA lesioning followed by chronic treatment with SCH-23390. Thus, bilateral destruction of dopamine-containing neurons and treatment with SCH-23390 in neonatal rats did not interfere with the developmental expression of D₁ receptors or alter the levels of mRNA that code for this receptor protein. Treatment of intact rats with SCH-23390 from postnatal day 4 to 20 also did not alter [³H]SCH-23390 binding or levels of D₁ receptor mRNA. However, adult rats treated chronically with SCH-23390 exhibited increased [³H]SCH-23390 binding but did not show a significant change in D₁ receptor mRNA levels.     

Involvement of Dopamine D1 and D2 Receptors in the Regulation of Proenkephalin mRNA Abundance in the Striatum and Accumbens of the Rat Brain

The effects of short-term treatment (6 h) with selective D1 or D2 agonists and antagonists on the mRNA for proenkephalin in the medial and anterior aspects of the caudate-putamen and the nucleus accumbens were assessed by in situ hybridization histochemistry. Proenkephalin mRNA abundance was significantly changed in the striatum and accumbens in response to D2 receptor manipulation. D2 blockade with haloperidol or raclopride increased, whereas D2 stimulation with LY-171555 (D2 agonist) decreased, striatal and accumbens proenkephalin mRNA abundance. Antagonism of D1 receptor activity with SCH-23390 significantly decreased proenkephalin mRNA abundance in all brain regions. Concurrent administration of the D1 agonist SKF-38393 prevented the SCH-23390 effect in all brain areas. The data demonstrate that acute treatment with dopaminergic D2 agonists and antagonists affects proenkephalin mRNA abundance in the striatum and accumbens via a D2 receptor mechanism, consistent with the concept that D2 receptor function inhibits the synthesis of the mRNA encoding the enkephalin peptides. Moreover, D1 receptor activity, directly or indirectly, exerts modulatory effects on proenkephalin mRNA abundance in the striatum and nucleus accumbens.     

Zinc Allosterically Modulates Antagonist Binding to Cloned D1 and D2 Dopamine Receptors

Abstract: Cations of various size and charge were used as atomic scale probes of D₁ and D₂ dopamine receptors. Those cations that perturbed the binding of D₁- and D₂-selective dopamine receptor antagonists were identified by screening at 5 m M cation. Pseudo-noble-gas-configuration d-transition metals, such as zinc, exerted a complete inhibition of specific binding, whereas most other cations had little or no effect. The nature of zinc's actions was characterized by measuring the radioligand binding properties of [³H]SCH-23390 and [³H]methylspiperone to cloned D_1A and D_2L dopamine receptors in either the presence or absence of Zn²⁺. Zinc exerts a low-affinity, dose-dependent, EDTA-reversible inhibition of the binding of subtype-specific antagonists primarily by decreasing the ligands' affinity for their receptors. The mechanism of zinc inhibition appears to be allosteric modulation of the dopamine receptor proteins because zinc increases the dissociation constant ( K _D) of ligand binding, Schild-type plots of zinc inhibition reach a plateau, and zinc accelerates antagonist dissociation rates. Here we demonstrate the effect of zinc on the binding of D₁- and D₂-selective antagonists to cloned dopamine receptors and show that the inhibition by zinc is through a dose-dependent, reversible, allosteric, two-state modulation of dopamine receptors.     

Pertussis Toxin Attenuates D2 Inhibition and Enhances D1 Stimulation of Adenylate Cyclase by Dopamine in Rat Striatum

The response of adenylate cyclase to GTP and to dopamine (DA) was investigated in synaptic plasma membranes isolated from rat striatum injected with pertussis toxin, which inactivates the inhibitory guanine nucleotide-binding regulatory protein (Ni) of adenylate cyclase. Pertussis toxin treatment reverted the inhibitory effects on the enzyme activity elicited by micromolar concentrations of GTP and reduced by 50% the DA inhibition of cyclase activity via D2 receptors. The toxin treatment enhanced the net stimulation of enzyme activity by DA in the presence of micromolar concentrations of GTP. However, the stimulatory effect of the selective D1 receptor agonist SKF 38393 was not significantly affected. The data indicate that Ni mediates D2 inhibition of striatal adenylate cyclase and participates in the modulation of D1 stimulation of the enzyme activity by DA.     

Dopamine D2 Receptor-Deficient Mice Exhibit Decreased Dopamine Transporter Function but No Changes in Dopamine Release in Dorsal Striatum

Abstract : Presynaptic D₂ dopamine (DA) autoreceptors, which are well known to modulate DA release, have recently been shown to regulate DA transporter (DAT) activity. To examine the effects of D₂ DA receptor deficiency on DA release and DAT activity in dorsal striatum, we used mice genetically engineered to have two (D₂^+/+), one (D₂^+/-), or no (D₂^-/-) functional copies of the gene coding for the D₂ DA receptor. In vivo microdialysis studies demonstrated that basal and K⁺-evoked extracellular DA concentrations were similar in all three genotypes. However, using in vivo electrochemistry, the D₂^-/- mice were found to have decreased DAT function, i.e., clearance of locally applied DA was decreased by 50% relative to that in D₂^+/+ mice. In D₂^+/+ mice, but not D₂^-/- mice, local application of the D₂-like receptor antagonist raclopride increased DA signal amplitude, indicating decreased DA clearance. Binding assays with the cocaine analogue [³H]WIN 35,428 showed no genotypic differences in either density or affinity of DAT binding sites in striatum or substantia nigra, indicating that the differences seen in DAT activity were not a result of decreased DAT expression. These results further strengthen the idea that the D₂ DA receptor subtype modulates activity of the striatal DAT.     

Methamphetamine-Induced Dopamine Overflow and Injury to Striatal Dopamine Terminals: Attenuation by Dopamine D1 or D2 Antagonists

Abstract: Pharmacological blockade of either D₁ or D₂ dopamine (DA) receptors prevents damage of striatal DA terminals by repeated doses of methamphetamine (m-AMPH). Because the substantial DA overflow produced by multiple m-AMPH treatments appears to contribute to the subsequent injury, we have investigated the effects of blockade of D₁ or D₂ receptors on m-AMPH-induced DA efflux using in vivo microdialysis. Four treatments with m-AMPH (4 mg/kg, s.c., 2-h intervals) produced large increases in striatal DA overflow, with particularly marked overflow (10 times the basal values) following the fourth injection. Administered by themselves, four injections of the D₁ antagonist SCH 23390 or the D₂ antagonist eticlopride (0.5 mg/kg, i.p., 2-h intervals) significantly increased striatal DA overflow. However, treatment with either SCH 23390 or eticlopride 15 min before each of four m-AMPH injections attenuated the marked DA peak otherwise seen after the fourth m-AMPH injection. These effects on DA overflow were related to subsequent DA depletions. Although our m-AMPH regimen produced a 54% reduction in striatal DA tissue content 1 week later, pretreatments with either the D₁ or the D₂ antagonist completely prevented subsequent DA content depletions. Furthermore, the DA content of striatal tissue remaining 1 week after m-AMPH treatment was significantly correlated with the magnitude of the cumulative DA overflow during the m-AMPH treatment ( r = -0.69). Thus, the extensive DA overflow seen during neurotoxic regimens of m-AMPH appears critical to the subsequent neurotoxicity, and the neuroprotective action of DA receptor antagonists seems to result from their attenuation of stimulant-induced DA overflow.     

Modulation of In Vivo Dopamine Release by D2 but Not D1 Receptor Agonists and Antagonists

The capacity of D1 and D2 agonists and antagonists to regulate the in vivo release and metabolism of dopamine (DA) in mesolimbic and nigrostriatal DA neurons of the mouse was determined using gas chromatographic and mass fragmentographic (GC-MF) analysis. DA release was inferred from levels of 3-methoxytyramine (3-MT) and DA metabolism was inferred from levels of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). DA release was increased by the D2 antagonists haloperidol and metoclopramide but not by the D1 antagonists SCH 23390 and SKF 83566. DA metabolism was increased by each of the four antagonists but to a greater extent with the D2 antagonists. The D2 agonists CGS 15855A and LY 171555 decreased DA release whereas the D1 agonist SKF 38393, at relatively high doses, only slightly affected DA release. Each of the three agonists decreased DA metabolism but again metabolism was more affected by the D2-selective drugs. The in vivo release of DA from mesolimbic and neostriatal DA neurons appears to be modulated by D2 but not by D1 receptors, whereas both receptor types can modulate DA metabolism.     

D2 Receptor Antagonists Do Not Modify Guanine Nucleotide-Sensitive Interactions Between Dopamine and D1 Dopamine Receptors Under In Vitro Conditions

Abstract: This study investigated possible D₁/D₂ interactions in rat and bovine striatal tissue by examining the effects of D₂ antagonists on the action of dopamine at D₁ dopamine receptors. In addition, the extent to which D₂ antagonists may induce an agonist low-affinity state of the D₁ receptor was evaluated in comparison with the effects of the guanine nucleotide analogue 5′-guanylylimidodiphosphate [Gpp(NH)p]. In saturation experiments dopamine caused a dose-dependent decrease in rat striatal and bovine caudate D₁ receptor density. This effect of dopamine, which has been shown to be sensitive to Gpp(NH)p, was not altered by pretreatment with either of the selective D₂ antagonists eticlopride (200 nM) or domperidone (200 nM). Results from displacement experiments show that the affinity of dopamine for D₁ receptors and the proportion of receptors in an agonist high-affinity state, are reduced by Gpp(NH)p (100 µM) but not by eticlopride. A molar excess of dopamine (100 µM) promotes the dissociation of (±)-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3-benzazepine-7-ol ([³H]SCH 23390) from rat striatal D₁ receptors at a rate that is significantly slower than when dissociation is initiated using 1 µM piflutixol. After pretreatment with Gpp(NH)p, [³H]SCH 23390 dissociation induced by dopamine occurred at an even slower rate. Pretreatment with eticlopride had no effect on the dopamine-induced rate of [³H]SCH 23390 dissociation. These results indicate that all experimental approaches detected dopamine effects at D₁ receptors that are Gpp(NH)p sensitive and D₂ antagonist insensitive and provide no evidence to support a D₁/D₂ link operating at the receptor level.     

D1 Dopamine Receptors of NS20Y Neuroblastoma Cells Are Functionally Similar to Rat Striatal D1 Receptors

Dopamine or agonists with D1 receptor potency stimulated cyclic AMP (cAMP) accumulation in whole cell preparations of NS20Y neuroblastoma cells. The accumulation of cAMP after D1 stimulation was rapid and linear for 3 min. Both dopamine and the novel D1 receptor agonist dihydrexidine stimulated cAMP accumulation two- to three-fold over baseline. The pseudo-Km for dopamine was approximately 2 microM, whereas for dihydrexidine it was approximately 30 nM. The effects of both drugs were blocked by either the D1-selective antagonist SCH23390 (Ki, 0.3 nM) or the nonselective antagonist (+)-butaclamol (Ki, 5 nM). Both (-)-butaclamol and the D2-selective antagonist (-)-sulpiride were ineffective (Ki greater than 3 microM). Forskolin (10 microM), prostaglandin E1 (1 microM), and adenosine (10 microM) also stimulated cAMP accumulation, but none were antagonized by SCH23390 (1 microM). Finally, muscarinic receptor stimulation (100 microM carbachol) inhibited both D1- and forskolin-stimulated increases in cAMP accumulation by 80%. The present results indicate that NS20Y neuroblastoma cells have D1 receptors that are coupled to adenylate cyclase, and that these receptors have a pharmacological profile similar to that of the D1 receptor(s) found in rat striatum.     

Dopamine Depletion Preferentially Impairs D1 over D2-Receptor Regulation of Striatal In Vivo Acetylcholine Release

The roles of D2 and D1 dopaminergic receptors on the regulation of striatal acetylcholine (ACh) release in vivo were examined for a period of 120 min after acute (2 h) or prolonged (16 h) depletion of brain dopamine (DA) by alpha-methyl-p-tyrosine. The reduction of DA transmission did not affect basal ACh output after 2 h but markedly lowered ACh release by 16 h (50%). Acute alpha-methyl-p-tyrosine pretreatment prevented the reduction of ACh release by the D1 antagonist SCH 23390 and its increase by the D2 antagonist, remoxipride, consistent with a drastic reduction of DA transmission at both DA receptors. However, 16 h after alpha-methyl-p-tyrosine, the effect of remoxipride on ACh release was restored, but SCH 23390 still had no effect, suggesting that the D2 inhibitory tone on ACh release had recovered, whereas the reduction of the D1 facilitatory influence persisted. The D1 facilitatory control of ACh neurotransmission thus appears to be more sensitive than the D2 inhibitory control to a reduction in DA transmission. The new model of DA-ACh interaction resulting from these data casts fresh light on the relationship between changes in DA transmission and extrapyramidal motor function.     

Effects of the Dopamine D3 Antagonist PD 58491 and Its Interaction with the Dopamine D3 Agonist PD 128907 on Brain Dopamine Synthesis in Rat

Abstract: The dopamine (DA) D₃ receptor antagonist PD 58491 {3-[4-[1-[4-[2-[4-(3-diethylaminopropoxy)phenyl]-benzoimidazol-1-yl-butyl]-1 H -benzoimidazol-2-yl]-phenoxy]propyl]diethylamine} bound with high affinity and selectivity to recombinant human DA D₃ versus D_2L and D_4.2 receptors transfected into Chinese hamster ovary cells: K _i values of 19.5 n M versus 2,362 and >3,000 n M , respectively. In contrast, the putative DA D₃ receptor antagonist (+)-AJ76 displayed low affinity and selectivity for D₃ versus D_2L and D_4.2 receptors (91 n M vs. 253 and 193 n M , respectively). In vitro, PD 58491 (1 n M −1µ M ) exhibited D₃ receptor antagonist activity, reversing the quinpirole (10 n M )-induced stimulation of [³H]thymidine uptake in D₃ CHOpro-5 cells, but did not have any significant intrinsic activity by itself in this assay. PD 58491 did not decrease the γ-butyrolactone-induced increase in DA synthesis ( l -3,4-dihydroxyphenylalanine accumulation) in rat striatum, indicating that the compound possessed no in vivo DA D₂/D₃ receptor agonist action at DA autoreceptors. PD 58491 (3–30 mg/kg, i.p.) generally did not alter DA or serotonin synthesis in either the striatum or mesolimbic region of rat brain. The D₃-preferring agonist PD 128907 decreased DA synthesis in striatum and mesolimbic regions, and this effect was attenuated by pretreatment with PD 58491. These findings support the hypothesis that DA D₃ autoreceptors may in part modulate the synthesis and release of DA in striatum and mesolimbic regions.     

Effects of Cerebral Ischemia on Regional Dopamine Release and D1 and D2 Receptors

Abstract: To expand on the nature of regional cerebral vulnerability to ischemia, the release of dopamine (DA) and dopaminergic (D₁ and D₂) receptors were investigated in Mongolian gerbils subjected to bilateral carotid artery occlusion (15 min) alone or with reflow (1–2 h). Extracellular cortical and striatal content of DA and its metabolites was measured by microdialysis using HPLC with electrochemical detection. The kinetic properties of D₁ and/or D₂ receptor binding sites were determined in cortical and striatal membranes with the use of radiolabeled ligands (¹²⁵I-SCH23982 and [³H]YM-09151-2, respectively). The ischemic release of DA from the striatum was greater (400-fold over preischemic level) than that from the cortex (12-fold over preischemic content). The affinity for the D₁-receptor ligand was lower ( K _D= 1.248 ± 0.047 n M ) after ischemia than that for sham controls ( K _D= 0.928 ± 0.032 n M, p < 0.001). The number of binding sites for D₂ receptors decreased in striatum ( B _max= 428 ± 18.4 fmol/mg of protein) after ischemia compared with sham controls ( B _max= 510 ± 25.2 fmol/mg of protein, p < 0.05). D₁ or D₂ binding sites were not changed either in the ischemic cortex or postischemic striatum and cortex. The findings strongly suggest that the ischemic release of DA from striatum is associated with early transient changes in D₁- and D₂-mediated DA neurotransmission.     

Binding of [3H]SCH23390 in Rat Brain: Regional Distribution and Effects of Assay Conditions and GTP Suggest Interactions at a D1-Like Dopamine Receptor

The compound [9-3H]SCH23390 [R-(+)-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3-benzazepine-7- ol] was synthesized, and the binding of this purportedly selective antagonist of D1 3,4-dihydroxyphenylethylamine (dopamine) receptors was characterized. The regional distribution of high-affinity, specific [3H]SCH23390 binding sites in the rat brain correlated well with levels of endogenous dopamine. Receptor densities were greatest in corpus striatum, nucleus accumbens, and olfactory tubercle; intermediate levels were found in several limbic and cortical areas, whereas few sites were detectable in cerebellum, brainstem, and ol-factory bulb. Specific binding in caudate-putamen was found to be both temperature- and pH-dependent, with optima at 25-30 degrees C and pH 7.8-8.0. Scatchard or Woolf analyses of binding in caudate-putamen suggest that most of the sites are either of a single class or of classes with similar characteristics (KD = 0.7 +/- 0.1 nM; Bmax = 347 +/- 35 fmol/mg of protein). Both dopamine and cis-flupenthixol altered the slope but not the intercept of lines generated by Scatchard analysis, suggesting a competitive mode of inhibition of [3H]SCH23390 binding. Competition for binding by dopamine or the D1 agonist SKF38393 was inhibited by guanine nucleotides, whereas GTP had little effect on the competition for binding by the antagonist cis-flupenthixol. The competition for [3H]SCH23390 binding sites by dopamine was much more sensitive to GTP than was competition for [3H]spiperone binding. These data support the hypotheses that [3H]SCH23390 binds to recognition sites that differ from those previously described using other radiolabeled dopamine antagonists and that these sites have the characteristics expected of dopamine receptors.     

Dopamine D1 and D2 Receptors and Their Signal System Present in Coated Vesicles Prepared from Bovine Striatal Tissue

Abstract: Coated vesicles (CVs) isolated from bovine striatal tissue were examined to determine whether they are associated with dopamine signal systems consisting of dopamine D₁ and D₂ receptors, G proteins, and adenylate cyclase. Dopamine receptors in CVs were characterized by a dopamine D₁ receptor antagonist, [³H]SCH 23390, and a dopamine D₂ receptor antagonist, [³H]-spiroperidol. The bindings of both ligands were specifically saturable and reversible with a dissociation constant ( K _D) of 0.65 and 0.5 n M , respectively. Dopaminergic antagonists and agonists inhibited the specific bindings of [³H]SCH 23390 and [³H]spiroperidol in a stereoselective and concentration-dependent manner with an appropriate rank order potency for dopamine D₁ or D₂ receptors. The regulations of the agonist binding by guanyl-5-ylimidodiphosphate were observed. ADP ribosylation of the CVs with [³²P]NAD demonstrated predominant labeling of bands of M_r 47,000–52,000, 42,000–45,000, and 40,000-39,000, which corresponded to the known molecular weights of the α subunits of G_s and G_i proteins. The presence of α and β subunits of G proteins in the CVs was also confirmed by immunoblotting assay. Adenylate cyclase activity, which was stimulated by SKF 38393 and inhibited by dopamine D₂ receptor agonists, was present in the CVs. These findings suggest that the dopamine D₁ and D₂ receptors in the CVs couple with adenylate cyclase via G_s/G_i protein.     

Endogenous Dopamine Facilitates Striatalln Vivo Acetylcholine Release by Acting on D1 Receptors Localized in the Striatum

Intrastriatal application of the D1 antagonist SCH 23390 by two procedures, reverse dialysis (20 microM) and local injection (0.45 nmol per striatum), elicited a reduction in acetylcholine (ACh) release superimposable on that induced by systemic administration. The novel selective D1 antagonist SCH 39166 produced a similar decreasing effect on striatal ACh release on local injection (0.45 nmol per striatum). On the other hand, local application of SCH 23390 into the frontal cortices (0.45 nmol per side) failed to alter striatal ACh overflow, indicating that the drug does not diffuse out of its injection site to any significant extent. The dopamine release inducer d-amphetamine (2 mg/kg s.c.) and the dopamine uptake inhibitor cocaine raised ACh release like the D1 agonists. These effects were completely blocked by 10 microM SCH 23390 applied by reverse dialysis. The results suggest that D1 receptors regulating ACh release are located in the striatum.     

Putative Neurotoxicity of SKF 38393 and Other D1 Dopaminergic Drugs Investigated in Rat Striatum

The recently alleged neurotoxicity of the D1 receptor agonist, SKF 38393, was investigated in rat striatum by measuring the enzymes acetylcholinesterase (AChE) and glutamate decarboxylase (GAD). First, unilateral intrastriatal microinjection of the excitotoxin kainic acid (2 micrograms in 1 microliter) was shown to evoke vigorous contraversive circling, followed 1 or 2 weeks later by profound decreases in striatal AChE (24 and 54%), GAD (51 and 75%), and protein (36 and 47%), as well as loss of GAD (45% at 2 weeks) in the ipsilateral substantia nigra. Similar striatal treatments with SKF 38393 (30 micrograms in 0.5-1 microliter), the related benzazepines SKF 82526 (D1 agonist, 30 micrograms in 1 microliter) and SCH 23390 (D1 antagonist, 5 micrograms in 1 microliter), or the phenanthridine D1 agonist CY 208-243 (5 micrograms in 1 microliter) failed to affect the rats' behaviour or their striatal levels of AChE, GAD, and protein. Intrastriatal SKF 38393 (30 micrograms in 0.5 microliter) also had no influence on these enzymes in the substantia nigra. It is concluded that none of the D1 dopaminergic compounds examined here was neurotoxic toward the many different cell groups that contain AChE and/or GAD in the striatum.     

Homologous Desensitization of the D1 Dopamine Receptor

Preincubation of D384 cells, derived from the human astrocytoma cell line G-CCM, with dopamine resulted in a time-dependent attenuation of cyclic AMP responsiveness to subsequent dopamine stimulation. This effect was agonist specific because the prostaglandin E1 (PGE1) stimulation of cyclic AMP of similarly treated cells remained unchanged. The attenuation by dopamine was concentration dependent with a maximum observed at 100 microM. A comparison of dopamine concentration-response curves of control and dopamine-preincubated cells revealed no change in the Ka apparent value, but a marked attenuation of the maximal response. Preincubation of cells with dopamine in the presence of D1 but not D2 selective antagonists partially prevented the observed attenuation. Attenuations in dopamine responsiveness were also obtained when D384 cells were preincubated with D1 but not D2 receptor agonists. The level of attenuation attained related to agonist efficiency in stimulating cyclic AMP: SKF38393 less than 3,4-dihydroxynomifensine less than fenoldopam less than 2-amino-6,7-dihydroxy-1,2,3,4-tetrahydronaphthalene = dopamine. However, increasing the efficiency of 3,4-dihydroxynomifensine stimulation of cyclic AMP, using the synergistic effect of adding a low concentration of forskolin, produced no further change in the attenuation of the subsequent response to dopamine. Thus, the D1 dopamine receptors expressed by D384 cells undergo homologous desensitization. Uncoupling of the D1 dopamine receptor appears to be independent of cyclic AMP formation, analogous to a mechanism proposed for the beta-adrenergic receptor.