D-1 dopamine receptor changes after striatal quinolinic acid lesion

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European Journal of Pharmacology, 138 (1987) 141-145

Elsevier EJP 224SC Short communication

D-1 dopamine receptor changes after striatal quinolinic acid lesion Paolo Barone, I d a Tucci, Sotirios A. Parashos a n d T h o m a s N. Chase * Experimental Therapeutics Branch, National Institute of Neurological and Communicative Disorders and Stroke, Building 10, Room 5C103, 9000 Rockoille Pike, Bethesda, MD 20892, U.S.A.

Received6 April 1987, accepted14 April 1987

Regional changes in the distribution of D-1 dopamine receptors produced by unilateral striatal lesions were evaluated in rat brain by quantitative autoradiography. Lesions were induced by local infusion of quinolinic acid, an endogenous excitotoxin that destroys intrinsic neurons, but spares fibers of passage. D-1 receptor density, as determined by [125I]SCH 23982 binding, was reduced in caudate-putamen, globus pallidus, substantia nigra, nucleus entopeduncularis on the lesioned side. These results confirm the presence of D-1 receptors on striatal cell bodies and provide direct evidence for the existence of presynaptic D-1 receptors on the terminals of striatal projections. D-1 dopamine receptors; [125I]SCH 23982; Quinolinic acid; Striatal lesion; Dopamine receptor interactions

1. Introduction

Two dopamine (DA) receptor subtypes, D-1 and D-2, are generally recognized by their contrasting effects on adenylate cyclase: D-1 receptor stimulation increases, while D-2 receptor stimulation inhibits the activity of this enzyme (Stool and Kebabian, 1981). These receptor subtypes have also been reported to mediate opposite effects on striatal DA metabolism (Sailer and Salama, 1985). Behavioral and neurophysiological studies, on the other hand, do not support such opposing actions: neither selective D-1 agonists nor selective D-2 agonists reverse reserpine-induced akinesia in mice, whereas the concomitant administration of both agonists restores locomotion (Gershanik et al., 1983). Concurrent D-1 and D-2 receptor stimulation also is required for the expression of DA-dependent postsynaptic effects on pallidal cell

* To whom all correspondence should be addressed: NINCDS/NIH, Building 10, Room 5C103, 9000 RockviUe Pike, Bethesda, MD 20892, U.S.A.

activity (Carlson et al., in press). The apparent inconsistency between the foregoing biochemical and behavioral-neurophysiological results might be attributable to differences in the distribution of D-1 and D-2 receptors: DA agonist-induced behaviors could result from effects at multiple, anatomically separated sites, whereas DA receptor-dependent adenylate cyclase production might reflect drug effects at a local, perhaps cellular, level. Previous reports from this laboratory have described the nature of D - l / D - 2 DA receptor interactions in the expression of motor function in rats with unilateral quinolinic acid-induced striatal lesions (Barone et al., 1986). D-1 receptor stimulation may provide 'tonic' activation, allowing D-2 receptor stimulation to become effective. In fact, ipsilateral turning was elicited by the selective D-2 agonist, LY 171555, but not by SKF 38393, a selective D-1 agonist. Blockade of D-1 receptors by SCH 23390, on the other hand, inhibited LY 171555-induced rotation, which could be restored by SKF 38393. Furthermore, when the selective agonists were given together, SKF 38393 aug-

0014-2999/87/$03.50 © 1987 Elsevier SciencePublishers B.V. (BiomedicalDivision)


mented LY 171555-induced turning (Barone et al., 1986). The present study sought an explanation for these behavioral findings based on the anatomic localization of dopamine receptor subtypes. We evaluated the distribution of D-1 dopamine receptors by means of quantitative autoradiography in rats with unilateral striatal lesions.

2. Materials and methods

2.1. Drugs Drugs used in this study included: quinolinic acid (Sigma Chemical Co., St. Louis, MO); SCH 23390 (Schering, Bloomfield, N J); SKF 38393 (Smith, Kline and French, Philadelphia, PA); LY 171555 (Eli Lilly and Co, Indianapolis, IN); [125I]SCH 23982, R(+)-81125I]iodo-7-hydroxy2,3,4,5-tetrahydro-3-methyl-5-phenyl-1 H-3-benzazepine, 2200 Ci/mmol (New England Nuclear, Boston, MA).

2.2. A nimals and autoradiographic procedures Male Sprague-Dawley rats (200 g) were anesthetized with pentobarbital (50 mg/kg i.p.) and placed in a David-Kopf stereotaxic apparatus. The left striatum was infused (coordinates according to Pellegrino-Cushman Atlas: A = 8.2, L = 2.5, V = 4.5) through a 29 gauge cannula for 2 rain with a 1/zl solution containing 300 nM of quinolinic acid dissolved in PBS at pH 7.4. The injection cannula was slowly removed 2 rain after the infusion. One week after lesioning, rats were killed under halothane anesthesia and perfused intracardially with cold 0.9% saline. Brains were rapidly removed, frozen by immersion into isopentane at - 7 0 ° C and sliced in 10 #m sections using a cryostat microtome set at - 1 7 ° C. Tissue sections were incubated for 30 rain at room temperature in coplin jars containing 50 mM Tris-HC1 buffer (pH 7.4) with the addition of 120 mM NaC1, 5 mM KC1, 2 mM CaCI2, 1 mM MgC12 and varying concentrations (0.05-2.5 nM) of [125I]SCH 23982. Non-specific binding was evaluated by incubating

serial sections under identical conditions with either 10 -6 M SCH 23390 or 50 nM ketanserin, the latter drug used to displace [125I]SCH 23982 binding to 5-HT2 receptors in selected brain areas. After incubation, tissues were washed twice for 5 min in buffer at 4 ° C and then briefly dipped into cold distilled water. In saturation experiments, tissue sections containing striatum were rinsed and then wiped from the slides using glass fiber filters. Bound radioactivity was determined by gamma counting. Specific binding of [125I]SCH 23982 was approximately 60-85% of the total binding. For autoradiogram generation, serial forebrain and brainstem sections that had been incubated in 0.1 nM [lzSI]SCH 23982 were rinsed, cold-air dried and placed on LKB-Ultrofilm (LKB Products, Rockville, MD) in Wolf X-ray cassettes for 16 h at 4°C. Images were analyzed using a series-68 (MTI, Japan) camera and computerized microdensitometry system (IBAS, C. Zeiss, West Germany). Optical densities of the autoradiograms were determined and converted to fmol of [125I]SCH 23982 bound per mg of protein using standards containing known quantities of [12sI]fl-endorphin that were exposed along with the brain sections. Protein content was determined by the Lowry method.

2.3. Data analysis Binding data obtained from either tissue sections or membrane suspensions were analyzed by the LIGAND program. Regional differences in D-1 receptor binding sites were evaluated by the two-tailed paired Student's t-test.

3. Results

[125I]SCH 23982 binding to striatal sections from brain-lesioned rats was saturable, specific and of high affinity. Computer analysis of the binding to tissue slices yielded a dissociation constant (K~) of 1.15 _ 0.1 nM and a maximal number of binding sites (Bmax) of 148 _+ 10 fmol/mg protein; the ligand bound to one site. In intact animals, [125I]SCH 23982 binding to tissue sec-

143 TABLE 1 Effect of unilateral striatal quinolinic acid injection on densities of [125I]SCH 23982 binding sites. Values reported are the means+S.E.M, of [125I]SCH 23982 specific binding for four rat brains. Specific binding was defined in tissue sections incubated in 0.1 nM [125I]SCH 23982 in absence and in presence of 10 -6 M SCH 23390 or 50 nM ketanserin. Brain region

[12sI]SCH 23982 specific binding fmol/mg of protein Per- SignifiUnlesioned Lesioned cent cance side side reduction

Fig. 1. Autoradiograms of two rat brain sagittal sections comparing D-1 receptor distribution on the quinolinic acid lesioned side (A) and the unlesioned side (B). SNR: substantia nigra pars reticulata, EP: entopeduncular nucleus, GP: globus pallidus, IC: internal capsule, Ch claustrum.

Accumbens Caudate-putamen Claustrum Cortex, lamina VI anterior cingulate entorhinal frontoparietal striate Entopeduncularn. Globus pallidus Internal capsule Olfactory tubercle Substantia nigra pars reticulata

58.4+ 6.0 72.5+12.0 18.5+ 6.3

55.9+6.9 4 6.2+0.7 91 16.4+2.7 11

n.s. P
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