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P Seeman

Publications and source records attributed to P Seeman.

At least 163 records · Page 9Linked to original sources
PubMed1985

Synthesis and radioreceptor binding activity of N-0437, a new, extremely potent and selective D2 dopamine receptor agonist.

The synthesis of a new, potent and selective D2 dopamine receptor agonist, N-0437, of the 2-aminotetralin group is described. The results of a radioreceptor binding assay using a homogenate of porcine anterior pituitary as a tissue source for D2 dopamine receptors and 3H-spiperone as radioligand demonstrate that this compound is one of the most potent compounds so far evaluated in this test system.

Animals↗
PubMed1985

Dependence of dopamine receptor conversion from agonist high- to low-affinity state on temperature and sodium ions.

Previous workers found that the anterior pituitary dopamine receptors were inconsistent in converting from their high-affinity state (D2 high) into their low-affinity state (D2 low) for dopamine. We tested, therefore, whether temperature or sodium ion concentration could be factors accounting for such inconsistencies. We found that the proportion of sites which converted depended on the temperature of incubation. No conversion occurred at 4 degrees, despite the presence of guanine nucleotide and sodium ions. At room temperature (20 degrees) guanine nucleotide consistently induced complete conversion in the presence of sodium ions, but gave inconsistent conversion in the absence of sodium ions. At body temperature (37 degrees) guanine nucleotide consistently resulted in complete conversion without requiring sodium ions.

Animals↗
PubMed1985

Dopamine D2 receptors photolabeled by iodo-azido-clebopride.

Iodo-azido-clebopride, a photoaffinity compound for dopamine D2 receptors, had high affinity for canine brain striatal dopamine D2 receptors with a dissociation constant (Kd) of 14 nM. Irradiation of striatal homogenate with iodo-azido-clebopride irreversibly inactivated 50% of dopamine D2 receptors at 20 nM (as indicated by subsequent [3H]spiperone binding). Dopamine agonists and antagonists prevented this photo-inactivation with the appropriate rank-order of potency. Striatal dopamine D1, serotonin (S2), alpha 1- and beta-adrenoceptors were not significantly inactivated following irradiation with iodo-azido-clebopride. Thus, iodo-azido-clebopride is a selective photoaffinity probe for dopamine D2 receptors, the radiolabelled form of which may aid in the molecular characterization of these proteins.

Affinity Labels↗
PubMed1985

Dopamine D2 receptors selectively labeled by a benzamide neuroleptic: [3H]-YM-09151-2.

In order to label dopamine D2 receptors selectively we tritiated the potent benzamide neuroleptic, YM-09151-2 (26.7 Ci/mmol). The binding of [3H]-YM-09151-2 to canine striatal membranes was saturable and specific with a KD of 57 pmol/l and Bmax of 36 pmol/g tissue as determined by Scatchard analysis. The KD, but not the Bmax, of [3H]-YM-09151-2 increased 6-fold in the absence of sodium chloride. [3H]-YM-09151-2 labeled 40% more sites than [3H]-spiperone in the same tissue homogenate. [3H]-YM-09151-2 binding was inhibited by dopaminergic drugs in a concentration and stereoselective manner with the appropriate dopamine D2 receptor profile. Thus, dopamine agonists inhibited [3H]-YM-09151-2 binding to canine striatal membranes with the following rank order of potency: (-)-N-n-propylnorapomorphine greater than apomorphine greater than (+/-)-6,7-dihydroxy-2-aminotetralin greater than (+)-N-n-propylnorapomorphine greater than dopamine greater than (-)-noradrenaline greater than serotonin greater than (-)-isoprenaline. Dopaminergic antagonists competed for [3H]-YM-09151-2 binding with the following order of potency: spiperone greater than (+)-butaclamol greater than haloperidol greater than clebopride greater than (-)-sulpiride greater than SCH-23390 greater than (-)-butaclamol. Furthermore, dopamine agonists recognized 2 states of the receptor labeled by [3H]-YM-09151-2, Dhigh2 and Dlow2. The Dhigh2 state of the receptor could be converted to Dlow2 by guanine nucleotides and sodium ions as is the case for [3H]-spiperone binding to D2 receptors. [3H]-YM-09151-2 appears to be a more selective ligand for dopamine D2 receptors than [3H]-spiperone, since YM-09151-2 displays approximately 9-fold lower affinity than spiperone for cortical serotonergic (S2) receptors. [3H]-YM-09151-2 may become a useful tool for the selective characterization of dopamine D2 receptors.

Animals↗
PubMed1985

L-dopa reverses the elevated density of D2 dopamine receptors in Parkinson's diseased striatum.

Striatal dopamine receptors were studied using [3H]-spiperone in postmortem tissues of thirty-six patients with Parkinson's Disease. Each tissue was analyzed by the receptor saturation method. In non-treated patients, the D2 dopamine receptor density was elevated in the caudate nucleus and putamen compared to controls. The dissociation constant for [3H]-spiperone was similar in all groups. The elevated density of D2 receptors in non-treated patients may indicate dopaminergic supersensitivity in this disease. The elevated density was reversed with dopamine agonist therapy, but the density was not lower than control tissues.

Adult↗
PubMed1985

Dopamine D2 receptors in brain and anterior pituitary recognize agonist and antagonist actions of (-)-3-PPP.

(-)-3-PPP, is a unique dopamine analogue, reported to have selective agonist actions at dopamine autoreceptors and antagonist actions at postsynaptic receptors. The interactions of D2 dopamine receptors with (-)-3-PPP in vitro were examined, using [3H]spiperone to label D2 receptors in brain regions containing both pre- and postsynaptic D2 receptors (caudate nucleus, corpus striatum) and a region containing nonsynaptic D2 receptors (anterior pituitary). In the absence of sodium ions, (-)-3-PPP detected D2 receptors in high- and low-affinity states in all regions examined, as is typical of dopamine agonists. That these two subpopulations of (-)-3-PPP-detected sites were dopaminergic in nature was assured by precluding [3H]spiperone binding to serotonergic receptors. In the presence of sodium ions, there was a significant increase in the affinity of some D2 receptors detected by (-)-3-PPP, and (-)-3-PPP in the presence of sodium was unable to discriminate between the two D2 affinity states in pituitary and striatum. The addition of guanine nucleotide led to (-)-3-PPP recognition of a single D2 binding site; the enhanced affinity of D2 receptors for (-)-3-PPP in the presence of sodium was retained in the presence of guanine nucleotide. These in vitro characteristics of (-)-3-PPP recognition of dopamine D2 receptor binding sites, when compared with dopamine and spiperone are seen to have clear features of both typical agonist and antagonist interactions with D2 receptors in both brain and pituitary.

Animals↗
PubMed1985

Complete conversion of brain D2 dopamine receptors from the high- to the low-affinity state for dopamine agonists, using sodium ions and guanine nucleotide.

Since previous work had shown that brain D2 3,4-dihydroxyphenylethylamine (dopamine) receptors were only partly converted from their high-affinity state to their low-affinity state, we here tested whether it was possible to obtain a complete 100% conversion of these receptors into their low-affinity state. It was first essential to resolve the components of [3H]spiperone binding to dopaminergic sites and nondopaminergic sites in rat striatal homogenates. In the presence of 50 microM S-sulpiride (to occlude the dopaminergic sites), therefore, we first determined that the residual binding of [3H]spiperone (approximately 20%) was inhibited by serotonergic agonists much more effectively than dopamine or noradrenaline, thus identifying the serotonergic component of [3H]spiperone binding. Thus, dopamine (or ADTN) inhibited the binding of [3H]spiperone at a high-affinity site (with dissociation constant of 10 nM dopamine), at a low-affinity site (with dissociation constant of 2,000 nM dopamine), and at the serotonergic site (with dissociation constant of 50,000 nM dopamine). In the absence of sodium ions, the high-affinity site was about 50% occupied by [3H]spiperone, and guanine nucleotide had no effect on this proportion. In the presence of 120 mM NaCl, however, the high-affinity site was reduced to 15% and guanine nucleotide completely eliminated this high-affinity site, 100% of the sites having been completely converted to their low-affinity state. Using [3H]N-propyl-norapomorphine to label the high-affinity state of the dopamine receptor, 50% conversion into the low-affinity state occurred at 45 mM LiCl, 69 mM NaCl, and 202 mM KCl. We conclude that it is possible to convert brain D2 dopamine receptors completely into their low-affinity state, in the presence of NaCl and a guanine nucleotide, providing that appropriate allowance is made for the serotonergic component of [3H]spiperone binding.

Animals↗
PubMed1985

Dopamine D2 receptors in the anterior pituitary: a single population without reciprocal antagonist/agonist states.

Although dopamine agonists can recognize two states of the D2 dopamine receptor in the anterior pituitary (D2high and D2low), we examined whether the dopamine antagonists such as [3H]spiperone could recognize these two sites with different affinities. Using up to 30 concentrations of [3H]spiperone, however, we could only detect a single population of binding sites (porcine anterior pituitary homogenates) with a dissociation constant (KD) of 130 pM. When specific [3H]spiperone binding was defined by a low concentration of (+)-butaclamol (100 nM), the apparent density was low. When defined by a high concentration of (+)-butaclamol (10 microM), nonspecific sites became detectable, thus revealing two apparent populations of sites for [3H]spiperone, only one of which was specific for dopamine. Sodium chloride reduced the KD of the single population of specific D2 sites to 64 pM. Guanine nucleotide by itself had no effect on the KD, but enhanced the density by 25%. Since the density-enhancement could be eliminated by extensive washing of membranes, and could be restored by preincubation with dopamine, the nucleotide-induced elevation of D2 density appeared to be a result of the release of tightly bound endogenous dopamine. Thus, monovalent cations and guanine nucleotides appear to have separate regulatory effects on the anterior pituitary D2 receptor that modulate antagonist-receptor interactions. Several maneuvers were used to test whether [3H]spiperone could differentiate between the two agonist-detected subpopulations of sites. Twentyfold different concentrations of [3H]spiperone (47 pM and 1000 pM) were found to label identical proportions of receptors in the D2high and D2low states as detected by the agonist 6,7-dihydroxyaminotetralin (ADTN), suggesting that spiperone labelled equal proportions of D2high and D2low sites without differential affinity for them. In addition, competition of spiperone for D2high sites selectively labelled by the agonist [3H]n-propylnorapomorphine (NPA) had a virtually identical KD for spiperone as did the total D2 receptor population as determined by direct binding studies (75 pM versus 64 pM). [3H]Spiperone also bound to a uniform population of D2low sites induced by preincubation with guanine nucleotide with identical affinity as to the total D2 population. Thus, these data do not support a "reciprocal model" for the D2 receptor (i.e., antagonist having low affinity for D2high and high affinity for D2low in a manner reciprocal to agonists).(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1985

Regulation of anterior pituitary D2 dopamine receptors by magnesium and sodium ions.

At D2 3,4-dihydroxyphenylethylamine (dopamine) receptors in anterior pituitary tissue, magnesium ions shifted receptors to agonist high-affinity states, but decreased the affinity of the antagonist [3H]spiperone. Conversely, sodium ions shifted the receptors to agonist low-affinity states, but increased the affinity of [3H]spiperone. Magnesium is proposed to stabilize the hormone-receptor-guanine nucleotide regulatory protein complex, whereas sodium appears to destabilize this ternary complex. Thus, magnesium and sodium appear to mediate their regulatory effects via a common component at the D2 dopamine-receptor ternary complex.

Animals↗
PubMed1985

Drug potencies on partially purified brain D2 dopamine receptors.

To examine the sensitivities of partially purified dopamine receptors to various dopaminergic agonists and antagonists, canine brain striatum dopamine receptors were enriched by isoelectric focusing. The digitonin-solubilized receptors were prelabelled with [3H]spiperone and focused for two time periods. After 5 h (incomplete focusing), radioactive peaks were detected at pH 6 and 9-11. Only the pH 6 peak revealed drug sensitivities expected of D2 receptors. Receptor recovery of the pH 6 peak was 79% with purification being sevenfold. After focusing overnight to equilibrium, the pH 6 peak further separated into peaks at pH 4.6 and 6.8. The receptor was identified only in the pH 4.6 fraction. The recovery of receptors in the pH 4.6 peak was low (10%), indicating little enrichment of the receptor. The rank order of binding of neuroleptics and dopamine agonists to the purified material was similar to that of the original preparation of soluble receptors. Dopamine did not bind to the purified pH 4.6 fraction unless the phosphate buffer (used during focusing) was replaced with Tris buffer. The absence of receptors in the pH 6.8 and pH 10 fractions, although both contained prelabeled [3H]spiperone, indicates the importance of testing agonists and antagonists on each fraction at each step in purification.

Animals↗
PubMed1985

The functional state of the dopamine receptor in the anterior pituitary is in the high affinity form.

In order to determine whether the high affinity state or the low affinity state of the dopamine receptor mediated the inhibition of release of PRL by dopamine agonists, a large number of dopaminergic agonists (n = 31) and antagonists (n = 24) were tested for their potencies to inhibit the binding of [3H] spiperone to porcine anterior pituitary tissue, and for their potencies to affect the release of PRL from rat anterior pituitary cells in culture. All agonists (except bromocriptine, ergocryptine, and dihydroergocryptine) inhibited [3H]spiperone binding in two phases: one phase occurred at nanomolar or subnanomolar concentrations (representing the high affinity state of the dopamine receptor) and the other phase occurred at much higher concentrations of agonist (the low affinity state of the dopamine receptor). The dissociation constants (K) for each drug at each state were derived by computer, with the program LIGAND. It was observed that the agonist K values for the high affinity state were virtually identical with those agonist concentrations inhibiting PRL release; the K values for the low affinity state were about 2 orders higher. These data suggest that the high affinity state of the D2 dopamine receptor is the functional state which mediates the inhibition of PRL release.

Animals↗
PubMed1985

Dopamine D2 receptor binding sites for agonists. A tetrahedral model.

In order to develop a model for the putative binding sites between the D2 dopamine receptor and many of its agonists, we obtained the dissociation constants of many dopaminergic agonists at the high affinity state, D2high, as well as at the low affinity state, D2low, of the receptor. [3H]Spiperone was used to label the D2 dopamine receptors in porcine anterior pituitary tissue. Agonists without any hydroxyl groups, such as 2-aminotetralin, effectively inhibited the binding of [3H]spiperone; the addition of a hydroxyl group corresponding to the "meta" position in dopamine, however, enhanced the potency (in four series of agonists) by an order of magnitude. The R-(-)-enantiomers of the aporphines and 5,6,-dihydroxy-2-dipropylaminotetralin were more potent than the S-(+)-enantiomers. Although the 4-methoxy-2-dipropylaminoindans were potent, the R-(-)-11-methoxyaporphines were not. A tetrahedral model is proposed; this has two sites for agonist attachment, the extremities of the sites being separated by 8 A, and their functional groups directed between 15 degrees and 30 degrees off the orthogonal from the receptor surface. Several steric obstacles are required to account for the inactivity of several congeners.

Animals↗
PubMed1985

Brain dopamine receptors in schizophrenia and tardive dyskinesia.

Brain dopamine receptors (type D2) mediate the psychomotor effects of dopamine. The D2 dopamine receptor can exist in either a high-affinity state for dopamine (nanomolar dissociation constant) or in a low-affinity state (micromolar dissociation constant). Both states of the receptor, however, have high affinity for neuroleptics (60 pM for spiperone). The postsynaptic receptor probably operates mainly in the D2 slow state. The presynaptic dopamine receptor, and also the dopamine receptors in the pituitary gland and the area postrema, probably function in the D2 high state. The density of brain D2 dopamine receptors is elevated in schizophrenia. The control densities were 10.5 pmol per g tissue. Half of the schizophrenic tissues (putamen, caudate nucleus, and nucleus accumbens) revealed densities of about 11.9 pmol per g, while the other half of the tissues revealed a density mode of 23.8 pmol per g. The bimodal distribution may support the concept of two types of schizophrenia. Future work must decide which group has more tardive dyskinesia.

Acetylcholine↗
PubMed1985

A photoaffinity ligand for dopamine D2 receptors: azidoclebopride.

In order to label D2 dopamine receptors selectively and covalently by means of a photosensitive compound, azidoclebopride was synthesized directly from clebopride. The dissociation constant (KD) of clebopride for the D2 dopamine receptor (canine brain striatum) was 1.5 nM, while that for azidoclebopride was 21 nM. The affinities of both clebopride and azidoclebopride were markedly reduced in the absence of sodium chloride. In the presence of ultraviolet light, azidoclebopride inactivated D2 dopamine receptors irreversibly, as indicated by the inability of the receptors to bind [3H]spiperone. Maximal photoinactivation of about 60% of the D2 dopamine receptors occurred at 1 microM azidoclebopride; 30% of the receptors were inactivated at 80 nM azidoclebopride (pseudo-IC50). Dopamine agonists selectively protected the D2 receptors from being inactivated by azidoclebopride, the order of potency being (-)-N-n-propylnorapomorphine greater than apomorphine greater than (+/-)-6,7-dihydroxy-2-aminotetralin greater than (+)-N-n-propylnorapomorphine greater than dopamine greater than noradrenaline greater than serotonin. Similarly, dopaminergic antagonists prevented the photoinactivation of D2 receptors by azidoclebopride with the following order of potency: spiperone greater than (+)-butaclamol greater than haloperidol greater than clebopride greater than (-)-sulpiride greater than (-)-butaclamol. The degree of D2 dopamine receptor photoinduced inactivation by azidoclebopride was not significantly affected by scavengers such as p-aminobenzoic acid and dithiothreitol. Furthermore, irradiation of striatal membranes with a concentration of azidoclebopride sufficient to inactivate dopamine D2 receptors by 60% did not significantly reduce dopamine D1, serotonin (S2), benzodiazepine, alpha 1- or beta-noradrenergic receptors. This study describes the use of a novel and selective photoaffinity ligand for brain dopamine D2 receptors. The molecule, in radiolabeled form, may aid in the molecular characterization of these receptors.

Affinity Labels↗