Lack of Dopaminergic Inputs Elongates the Primary Cilia of Striatal Neurons

In the rodent brain, certain G protein-coupled receptors and adenylyl cyclase type 3 are known to localize to the neuronal primary cilium, a primitive sensory organelle protruding singly from almost all neurons. A recent chemical screening study demonstrated that many compounds targeting dopamine receptors regulate the assembly of Chlamydomonas reinhardtii flagella, structures which are analogous to vertebrate cilia. Here we investigated the effects of dopaminergic inputs loss on the architecture of neuronal primary cilia in the rodent striatum, a brain region that receives major dopaminergic projections from the midbrain. We first analyzed the lengths of neuronal cilia in the dorsolateral striatum of hemi-parkinsonian rats with unilateral lesions of the nigrostriatal dopamine pathway. In these rats, the striatal neuronal cilia were significantly longer on the lesioned side than on the non-lesioned side. In mice, the repeated injection of reserpine, a dopamine-depleting agent, elongated neuronal cilia in the striatum. The combined administration of agonists for dopamine receptor type 2 (D2) with reserpine attenuated the elongation of striatal neuronal cilia. Repeated treatment with an antagonist of D2, but not of dopamine receptor type 1 (D1), elongated the striatal neuronal cilia. In addition, D2-null mice displayed longer neuronal cilia in the striatum compared to wild-type controls. Reserpine treatment elongated the striatal neuronal cilia in D1-null mice but not in D2-null mice. Repeated treatment with a D2 agonist suppressed the elongation of striatal neuronal cilia on the lesioned side of hemi-parkinsonian rats. These results suggest that the elongation of striatal neuronal cilia following the lack of dopaminergic inputs is attributable to the absence of dopaminergic transmission via D2 receptors. Our results provide the first evidence that the length of neuronal cilia can be modified by the lack of a neurotransmitter''s input.     

Effect of long-term L-dopa administration on the dopaminergic and cholinergic (muscarinic) receptors of striatum in 6-hydroxydopamine lesioned rats

L-Dihydroxyphenylalanine (L-Dopa) (200 mg/kg/day) was administered for 30 days to the rats whose nigrostriatal dopamine pathway was lesioned unilaterally with 6-hydroxydopamine and the receptor binding of ³H-spiperone and ³H-quinuclidinyl benzilate (³HQNB) was measured in the dopaminergic and muscarinic cholinergic receptors of the striatum. ³H-spiperone binding increased by 73% and ³HQNB binding decreased by 14% in the lesioned side when compared to the control side of L-Dopa-non-treated rats. ³H-spiperone binding was measured in the lesioned sides of L-Dopa-treated and L-Dopa-non-treated rats and was found to have decreased by 21% in the former. In the control side of the L-Dopa-treated lesioned rats, however, ³H-spiperone binding increased by 27% when compared to the opposite striatum of the same rats. ³HQNB binding in the lesioned side of L-Dopa-treated rats was not significantly different from that of the control side statistically. These results suggest that changes in functional equilibrium between the dopaminergic and cholinergic mechanisms influence the muscarinic cholinergic receptors and that supersensitivity of dopamine receptors after lesion of the nigrostriatal pathway also remains after long-term L-Dopa treatment.     

Effects of unilateral 6-OHDA lesions on [3H]-N-propylnorapomorphine binding in striatum ex vivo and vulnerability to amphetamine-evoked dopamine release in rat

It has been argued that agonist ligands for dopamine D(2/3) receptors recognize a privileged subset of the receptors in living striatum, those which are functionally coupled to intracellular G-proteins. In support of this claim, the D(2/3) agonist [(3)H]-N-propylnorapomorphine ([(3)H]NPA) proved to be more vulnerable to competition from endogenous dopamine than was the antagonist ligand [(11)C]raclopride, measured ex vivo in mouse striatum, and subsequently in multi-tracer PET studies of analogous design. Based on these results, we predicted that prolonged dopamine depletion would result in a preferential increase in agonist binding, and a lesser competition from residual dopamine to the agonist binding. To test this hypothesis we used autoradiography to measure [(3)H]NPA and [(3)H]raclopride binding sites in hemi-parkinsonian rats with unilateral 6-OHDA lesions, with and without amphetamine challenge. Unilateral lesions were associated with a more distinct increase in [(3)H]NPA binding ex vivo than was seen for [(3)H]raclopride binding in vitro. Furthermore, this preferential asymmetry in [(3)H]NPA binding was more pronounced in amphetamine treated rats. We consequently predict that agonist ligands should likewise be fitter than antagonists for detecting responses to denervation in positron emission tomography studies of idiopathic Parkinson's disease. Agonist binding increases in vivo are likely to reflect the composite of a sensitization-like phenomenon, and relatively less competition from endogenous dopamine, as seen in the lesioned side of 6-OHDA induced hemi-parkinsonism.     

In vivo binding of 125I-LSD to serotonin 5-HT2 receptors in mouse brain

The binding of 125I-LSD (2-[125I]-lysergic acid diethylamide) was studied in various mouse brain regions following intravenous injection of the radioligand. The high specific activity of 125I-LSD enabled the injection of low mass doses (14 ng/kg), which are well below the threshold for induction of any known physiological effect of the probe. The highest levels of 125I-LSD binding were found in the frontal cortex, olfactory tubercles, extra-frontal cortex and striatum while the lowest level was found in the cerebellum. Binding was saturable in the frontal cortex but increased linearly in the cerebellum with increasing doses of 125I-LSD. Serotonergic compounds potently inhibited 125I-LSD binding in cortical regions, olfactory tubercles, and hypothalamus but had no effect in the cerebellum. Dopaminergic compounds caused partial inhibition of binding in the striatum while adrenergic compounds were inactive. From these studies we conclude that 125I-LSD labels serotonin 5-HT2 receptor sites in cortical regions with no indication that other receptor sites are labeled. In the olfactory tubercles and hypothalamus, 125I-LSD labeling occurs predominantly or entirely at serotonin 5-HT2 sites. In the striatum, 125I-LSD labels approximately equal proportions of serotonergic and dopaminergic sites. This data indicates that 125I-LSD labels serotonin receptors in vivo and suggests that appropriate derivatives of 2I-LSD may prove useful for tomographic imaging of serotonin 5-HT2 receptors in the mammalian cortex.     

125I](+/-)FISCH: a new CNS D-1 dopamine receptor imaging ligand

Radiolabeling and in vitro and in vivo evaluation of an iodinated benzazepine: [125I] FISCH 7-Chloro-8-hydroxy-1-(4'-iodophenyl)-3-methyl-2,3,4,5- tetrahydro-1H-3-benzazepine, as a potential imaging agent for CNS D-1 dopamine receptors in animals, were investigated. After an iv injection, this benzazepine derivative showed good brain uptake in rats (2.70, 1.28, 0.48 %dose/whole brain at 2, 15 and 60 min, respectively). The striatum/cerebellum ratio was 2.50 at 60 min after the injection. The regional distribution in rat brain, as measured by ex vivo autoradiography, displayed highest uptake in the regions of the striatal complex and the substantia nigra, regions known to have a high concentration of D-1 dopamine receptors. Furthermore, this localized regional cerebral distribution was blocked by pretreatment with SCH-23390, a selective D-1 dopamine receptor antagonist. The in vitro binding affinity of this agent in rat striatum tissue preparation displayed a Kd of 1.43 +/- 0.15 nM. Competition data (in vitro) showed the following rank order of potency: SCH-23390 greater than (+/-)IBZP much greater than apomorphine greater than WB 4101 greater than ketanserin approximately spiperone. The preliminary data suggest that this analog of SCH-23390 shows similar selectivity for the CNS D-1 receptor.     

Effect of Unilateral Perinatal Hypoxic-Ischemic Brain Injury in the Rat on Dopamine D1 and D2 Receptors and Uptake Sites: A Quantitative Autoradiographic Study

The effect of a unilateral perinatal hypoxic-ischemic brain injury on dopamine D1 and D2 receptors and uptake sites was investigated in rats by using in vitro quantitative binding autoradiography, 2-3 weeks after the insult. We observed significant decreases in the Bmax and KD for [3H]SCH 23390-labeled D1 and in the Bmax for [3H]spiperone-labeled D2 receptors in the lesioned caudate-putamen in rats with moderate brain injury (visible loss in hemispheric volume ipsilateral to the injury) compared with the nonlesioned contralateral caudate-putamen or with control rats. Changes in [3H]SCH 23390 and [3H]spiperone binding predominated in the dorsolateral part of the lesioned caudate-putamen. Pronounced reduction in [3H]SCH 23390 binding was also observed in the substantia nigra pars reticulata on the side of the lesion. In contrast, we did not observe any significant change in Bmax or KD for [3H]mazindol-labeled dopamine uptake sites. Similarly, no significant changes in the levels of dopamine or its metabolites were found on the side of the lesion. The observed reductions in striatal dopamine D1 and D2 receptors are a reflection of striatal cell loss induced by the hypoxic-ischemic injury. The absence of changes in [3H]mazindol binding or dopamine levels in the lesioned caudate-putamen indicates that the dopaminergic presynaptic structures are preserved.     

5-Iodo-A-85380 binds to alpha-conotoxin MII-sensitive nicotinic acetylcholine receptors (nAChRs) as well as alpha4beta2* subtypes

Recent work suggests that 5-iodo-A-85380, a radioiodinated analog of the 3-pyridyl ether A-85380, represents a promising imaging agent for non-invasive, in vivo studies of alphaAbeta2* nicotinic acetylcholine receptors (nAChRs; *denotes receptors containing the indicated subunits), because of its low non-specific binding, low in vivo toxicity and high selectivity for alpha4beta2* nAChRs. As an approach to elucidate nAChR subtypes expressed in striatum, we carried out competitive autoradiography in monkey and rat brain using 5-[125I]iodo-A-85380 ([125I]A-85380) and [125I]alpha-conotoxin MII, a ligand that binds with high affinity to alpha6* and alpha3* nAChRs, but not to alpha4beta2* nAChRs. Although A-85380 is reported to be selective for alpha4beta2* nAChRs, we observed that A-85380 completely inhibited [125I]alpha-conotoxin MII binding in rat striatum and that A-85380 blocked >90% of [125I] alpha-conotoxin MII sites in monkey caudate and putamen. These results suggest that A-85380 binds to non-alpha4beta2* nAChRs, including putative alpha6* nAChRs. Experiments to determine the percentage of [125I]A-85380 sites that contain alpha-conotoxin MII-sensitive (alpha6beta2*) nAChRs indicate that they represent about 10% of [125I]A-85380 sites in rodent striatum and about 30% of sites in monkey caudate and putamen. These data are important for identifying alterations in nicotinic receptor subtypes in Parkinson's disease and other basal ganglia disorders both in in vitro and in in vivo imaging studies.     

Radioiodinated D-(+)-N1-ethyl-2-iodolysergic acid diethylamide: a ligand for in vitro and in vivo studies of serotonin receptors

Radioiodinated D-(+)-N1-ethyl-2-iodolysergic acid diethylamide ([125I]-EIL) has been evaluated as a ligand for in vitro and in vivo studies of cerebral serotonin 5-HT2 receptors. [125I]-EIL exhibited high affinity (KD = 209 pM) for 5-HT2 receptors with a high degree of specific binding (80-95%) in membranes from rat prefrontal cortex. The regional distribution of [125I]-EIL binding in vivo to seven areas of mouse brain correlated significantly (Rs = 0.93) with known densities of 5-HT2 receptors. In vivo specificity, defined by tissue to cerebellum radioactivity ratios, reached a maximum for frontal cortex at 6 hr (21.2) and persisted through 16 hr (8.8). Ketanserin, a 5-HT2 receptor antagonist, fully inhibited binding in a dose dependent fashion in all brain regions except cerebellum. By contrast, blockers for dopamine D2, alpha- or beta-adrenergic receptors did not significantly inhibit radioligand binding in any region. [125I]-EIL selectively labels 5-HT2 receptors in vivo with the highest specificity of any serotonergic ligand reported to date, indicating that [123I]-EIL should prove applicable to single photon emission computed tomography studies in living brain.     

Characterization of Radioiodinated TISCH: A High-Affinity and Selective Ligand for Mapping CNS D1 Dopamine Receptor

In developing CNS D1 dopamine receptor-imaging agents with improved specificity and longer brain retention, an iodinated D1 ligand was synthesized. In vitro and in vivo radiolabeling studies of a new iodinated benzazepine, TISCH [7-chloro-8-hydroxy-1-(3'-iodophenyl)-3-methyl-2,3,4,5-tetrahydro-1H-3- benzazepine], an analog of SCH 23390 (7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepin e), were investigated. After an intravenous injection, the R(+) isomer of TISCH showed high brain uptake in rats (2.20 and 0.57% dose per whole brain at 2 and 60 min, respectively). The striatum/cerebellum ratio increased progressively with time (12 at 60 min). Ex vivo autoradiography of rat brain sections, after intravenous injection of R(+)-[125I]TISCH, displayed the highest uptake in striatum and substantia nigra, regions known to have a high concentration of D1 receptors, whereas the S(-) isomer displayed no specific uptake. Furthermore, the specific uptake can be blocked by pretreatment with SCH 23390. In vitro binding studies using the rat striatum tissue preparation showed high specific and low nonspecific bindings (KD = 0.21 +/- 0.03 nM). The rank order of potency exhibiting high specificity to the D1 receptor was SCH 23390 greater than (+/-)-TISCH greater than (+)-butaclamol = (+/-)-FISCH [7-chloro-8-hydroxy-1-(4'-iodophenyl)-3-methyl-2,3,4,5-tetrahydro-1 H-3-benzazepine] much greater than WB4101 = spiperone greater than dopamine, serotonin, (+/-)-propranolol, and naloxone. Imaging studies in a monkey with the resolved isomer, R(+)-[123I]TISCH, demonstrated a high uptake in the basal ganglia and prolonged retention. The preliminary data suggest that R(+)-TISCH is selective for the CNS D1 receptor and is potentially useful for in vivo and in vitro pharmacological studies. When labeled with iodine-123, it may be suitable for noninvasive imaging in humans.     

Changes in striatal specific 3-H-atropine binding after unilateral 6-hydroxydopamine lesions of nigrostriatal dopaminergic neurones.

Specific binding of ³H-atropine to crude synaptosomal membrane fractions of the rat striatum was measured at different times after unilateral 6-hydroxydopamine lesions of the nigrostriatal dopaminergic neurones. In a group of rats killed between 4 to 15 days after lesioning the right side, specific ³H-atropine binding was reduced by 20 percent compared to the right side of unlesioned rats. There was a concomitant increase (20 percent) of specific ³H-atropine binding in the contralateral side compared to control animals. These changes in muscarinic receptor binding depended on the time after which the lesions were made : maximum effects occured about 8 days after lesioning but almost completely disappeared 13 days later. Dissociation constants for ³H-atropine in the right and left striata of control and lesioned rats were not significantly different. The decrease in muscarinic receptor binding in the ipsilateral striatum of lesioned animals may result from an activation of cholinergic neurones produced by removal of the inhibitory dopaminergic terminals.     

Comparison of In vivo D-2 dopamine receptor binding of IBZM and NMSP in rat brain

In vivo biodistribution of S- and R-isomers of [¹²⁵I]IBZM in rats showed a significant initial brain uptake (3.20 and 2.67% dose/organ at 2 min, respectively). The wash-out from the brain was slower for the S-isomer. The striatum to cerebellum ratio for [¹²⁵I]S-IBZM decreased with an increasing dose of cold carrier or spiperone, suggesting that the brain uptake is stereospecific and saturable, and may be related to the binding of D-2 dopamine receptors. In a dual isotope digital autoradiography study [¹²⁵I]IBZM and [³H]NMSP(N-methylspiperone) show comparable regional cerebral distribution in rats.     

Pharmacological evaluation of (+)-2- [123I]A-69024: a radioligand for in vivo studies of dopamine D1 receptors

(+)-2-[123I] A-69024, [(+)-1-(2-[123I] iodo-4,5-dimethoxy-benzyl)-7-hydroxy-6-methoxy-2-methyl-1,2,3,4-tetrahydroisoquinoline], is a specific and enantioselective dopamine D1 receptor radioligand. The in vivo biodistribution of this radioligand in rats showed high brain uptake and a distribution consistent with the density of dopamine D1 receptors. Highest uptake was observed in the striatum (0.65 %ID/g) at 5 min followed by clearance. As a measure of specificity the striatum/cerebellar ratio reached a maximum of 3.9 at 30 min post-injection. Radioactivity in the striatum was reduced by 68% by pre-administration of the D1 antagonist SCH 23390. Pre-administration of other dopamine binding drugs, spiperone (D2), 7-OH-DPAT (D3), and clozapine (D4) displayed no inhibitory effect on (+)-2-[123I]A-69024 accumulation in any brain region. Ketanserin (5-HT2/5-HT2C) and haloperidol (D2 receptor antagonist/sigma receptor ligand) also displayed no inhibitory effect in any brain region studied. With the pharmacologically inactive enantiomer, (-)-2-[123I]A-69024, the brain uptake was determined to be non-specific since a striatum/cerebellar ratio of near 1 was observed throughout the time course of the experiment. (+)-2-[123I]A-69024 displays enantioselectivity for dopamine D1 receptors and may deserve further investigation as a possible SPECT radioligand.     

No change in dopamine D1 receptor in vivo binding in rats after sub-chronic haloperidol treatment

A frequent side effect in the long-term treatment of schizophrenia with the dopamine D2 antagonist haloperidol (HAL) is the appearance of tardive dyskinesia or, in animals, of repetitive involuntary vacuous chewing movements (VCMs). In rats, chronic HAL-induced or D1 receptor-stimulated VCMs are suppressed by D1 antagonists, suggesting that this behavioral supersensitivity is mediated by D1 receptors. The goal of this study was to investigate in vivo the possible relationship between D1 receptor binding and D1-mediated behavioral supersensitivity, after subchronic HAL treatments. D1 agonist R-SKF 82957 and antagonist SCH 23390, both labeled with carbon-11, were used to assess in vivo D1 receptor binding. Rats were treated with HAL (1.5 mg/kg, i.p.) or vehicle for 21 days, followed by a 4 day washout period. No significant difference was found in the regional brain binding of either radioligand. D1 receptor-mediated behaviors including VCMs, grooming, and rearing were measured in control or HAL-treated rats. VCMs were significantly increased in HAL-treated rats, suggesting D1 receptor stimulation and possibly receptor supersensitivity. This study failed to link the purported D1 receptor-mediated behaviors with in vivo receptor binding measures of R-[11C]SKF 82957 or [11C]SCH 23390 in rat brain regions.     

Iodoethylspiperone,a new potential agent for exploration of central dopamine D2 receptors: Synthesis and preliminary in vivo study

We synthesized a new spiperone derivative: iodoethylspiperone (IES) to perform dopamine D₂ receptor exploration by SPECT. IES was prepared from a precursor: tosylethylspiperone, and characterized by i.r. and ¹H-NMR analyses. [¹²⁵I]IES was obtained with 80% yield. In vivo biodistribution in rats showed a high and specific uptake in the striatum. The uptake ratio between the striatum and the cerebellum reached a maximum value 4 h after injection (10.05 ± 2.81). IES labeled with ¹²³I should be a promising new agent to investigate D₂ receptors in the living human brain.     

Radioiodinated 2'-iodospiperone: a new radioligand for in vivo dopamine receptor study

In vivo dopamine receptor binding of the newly synthesized ligand, 125I-2'-iodospiperone (125I-2'-ISP), was studied in mouse brain. The highest accumulation was found in the striatum. Analysis of the striatal homogenate showed the 125I-2'-ISP to be metabolically stable. Furthermore, this striatal binding was saturable and displaced only by dopaminergic drugs. On the other hand, the accumulation in the cortex was as low as that of the cerebellum and uneffected by the administration of serotoninergic drugs and dopaminergic drugs; results assessed by macroautoradiographic studies. Thus, the newly synthesized 125I-2'-ISP presented high affinity for dopamine receptors in vivo and therefore, holds great potential for the in vivo dopamine receptor studies, provided 123I becomes readily available.     

High affinity dopamine D2 receptor radioligands. 2. [125I]epidepride, a potent and specific radioligand for the characterization of striatal and extrastriatal dopamine D2 receptors.

Epidepride, (S)-N-[(1-ethyl-2-pyrrolidinyl)methyl]-5-iodo-2,3-dimethoxybenzamide+ ++, the iodine analogue of isoremoxipride (FLB 457), was found to be a very potent dopamine D2 receptor antagonist. Optimal in vitro binding required incubation at 25 degrees C for 4 h at pH 7.4 in a buffer containing 120 mM NaCl, 5 mM KCl, 2 mM CaCl2 and 1 mM MgCl2. Scatchard analysis of in vitro binding to striatal, medial frontal cortical, hippocampal and cerebellar membranes revealed a KD of 24 pM in all regions, with Bmax's of 36.7, 1.04, 0.85, and 0.37 pmol/g tissue, respectively. The Hill coefficients ranged from 0.91-1.00 in all four regions. The IC50's for inhibition of [125I]epidepride binding to striatal, medial frontal cortical, and hippocampal membranes for SCH 23390, SKF 83566, serotonin, ketanserin, mianserin, naloxone, QNB, prasozin, clonidine, alprenolol, and norepinephrine ranged from 1 microM to greater than 10 microM. Partial displacement of [125I]epidepride by nanomolar concentrations of clonidine was noted in the frontal cortex and hippocampus, but not in the striatum. Scatchard analysis of epidepride binding to alpha 2 noradrenergic receptors in the frontal cortex and hippocampus revealed an apparent KD of 9 nM. At an epidepride concentration equal to the KD for the D2 receptor, i.e. 25 pM, no striatal alpha 2 binding was seen and only 7% of the specific epidepride binding in the cortex or hippocampus was due to binding at the alpha 2 site. Correlation of inhibition of [3H]spiperone and [125I]epidepride binding to striatal membranes by a variety of D2 ligands revealed a correlation coefficient of 0.99, indicating that epidepride labels a D2 site. In vitro autoradiography revealed high densities of receptor binding in layers V and VI of prefrontal and cingulate cortices as well as in striatum. In vivo rat brain uptake revealed a hippocampal:cerebellar and frontal cortical:cerebellar ratio of 2.2:1 which fell to 1.1:1 following haloperidol pretreatment. These properties suggest that [125I]epidepride is a superior radioligand for the in vitro and in vivo study of striatal and extrastriatal dopamine D2 receptors.     

Effect of desipramine on dopamine receptor binding in vivo

Effect of desipramine (given i.p. 30 min prior to the tracer injection) on the in vivo binding of 3H-SCH23390 and 3H-N-methylspiperone (3H-NMSP) in mouse striatum was studied. The ratio of radioactivity in the striatum to that in the cerebellum at 15 min after i.v. injection of 3H-SCH23390 or 45 min after injection of 3H-NMSP were used as indices of dopamine D1 or D2 receptor binding in vivo, respectively. In vivo binding of D1 and D2 receptors was decreased in a dose-dependent manner by acute treatment with desipramine (DMI). A saturation experiment suggested that the DMI-induced reduction in the binding was mainly due to the decrease in the affinity of both receptors. No direct interactions between the dopamine receptors and DMI were observed in vitro by the addition of 1 mM of DMI into striatal homogenate. Other antidepressants such as imipramine, clomipramine, maprotiline and mianserin also decreased the binding of dopamine D1 and D2 receptors. The results indicated an important role of dopamine receptors in the pharmacological effect of antidepressants.     

Intraventricular infusion of leupeptin decreases Bmax of the D2 receptor in the striatum of young rats.

Intraventricular infusion of a thiol protease inhibitor, leupeptin, was previously shown to induce several morphological and immunochemical manifestations of normal and pathological aging in rat brain. The present study attempted to elucidate whether this treatment also perturbs another brain function which declines in aging, dopamine D2 receptor binding in striatum. Intraventricular infusion of leupeptin (0.6 mg per day) for two weeks caused a significant (about 20%) reduction in the binding maximum (Bmax) of dopamine D2 receptors (as examined by [3H] spiperone binding) in the striatum of young male Fischer-344 rats in comparison to (saline-infused) control rats. The apparent Kd values did not differ significantly between the control and leupeptin-treated rat groups. The results suggest that decreased protein turnover may be a factor in the decline in Bmax of D2 receptors during aging.     

In vitro binding properties and autoradiographic imaging of 3-iodobenzamide ([125I]-IBZM): a potential imaging ligand for D-2 dopamine receptors in SPECT

The in vitro binding properties of the [125I] labeled benzamide (S(-)-N-[(1-ethyl-2-pyrrolidinyl)-methyl]-2-hydroxy-3-iodo-6-methoxy- benzamide, IBZM) were determined in bovine and mouse caudate membrane homogenates and by autoradiography of mouse brain slices. [125I]-IBZM binding is saturable and reversible with a Bmax of 373 +/- 51 fmol/mg protein and a Kd of 3.1 +/- 0.62 nM (mean +/- SD, Scatchard analyses) and 0.56 nM as calculated by association and dissociation time constants. In competition experiments, Ki values for the D-2 antagonists YM-09151-2 and spiperone are 4 orders of magnitude lower than the Ki value for the D-1 antagonist SCH-23390 and S(-)-IBZM is ten-fold more potent than R(+)-IBZM. [125I]-IBZM has a low affinity for serotonin S-2 and for alpha receptors. Therefore, it is a highly selective ligand for dopamine D-2 receptors. Autoradiographic images of brain sections incubated with [125I]-IBZM show the dopamine D-2 receptors of the striatum, nucleus accumbens and olfactory tubercle with a high ratio of specific to nonspecific binding. Thus, S(-)-IBZM, when labeled with [123I], may be useful for in vivo imaging of dopamine D-2 receptors by single photon emission computerized tomography (SPECT).