Pierre Demenge

Identification and Distribution of Neuroleptic Binding Sites in the Rat Spinal Cord

Abstract: Identification of neuroleptic receptor sites in the rat spinal cord could be achieved by the binding of [3H]haloperidol to membranes taken from the different horns. The use of pooled frozen microdiscs punched from these different spinal cord areas allowed the detection of saturable stereospecific binding, as defined in the presence of (+)‐ and (‐)‐butaclamol. Comparison of the binding constants with those obtained in the corpus striatum resulted in similar dissociation constants and Hills slopes. Maximal binding capacity was quite different, being the greatest in the whole striatum (157 ± 8 fmol/mg protein) followed by the dorsal horn (56 ± 3 fmol/mg protein) and the lateral (34 ± 5 fmol/mg protein) and ventral ones (31 ± 2 fmol/mg protein). The displacement of the labelled ligand by different dopaminergic and nondopaminergic drugs at various concentrations gave similar results in the whole striatum and the spinal cord, giving further support for the existence of a dopaminergic innervation of the spinal cord and showing that dopaminergic receptor sites are distributed through the different spinal horns, with a maximal density in the dorsal horn–as for dopamine levels. No detectable stereospecific binding could be obtained from the surrounding spinal white matter, even at high tissue concentrations. Owing to poor sensitivity of the binding technique, no stereospecific neuroleptic binding could be demonstrated in the whole spinal cord, even at very high tissue concentration, whereas it could be detected in spinal cord tissue sampled from restricted areas of dense dopaminergic innervation.

Supersensitivity time course of dopamine antagonist binding after nigrostriatal denervation: Evidence for early and drastic changes in the rat corpus striatum

The sensitivity of the [3H]haloperidol binding technique can be greatly increased by focusing tissue sampling on striatal regions where dopaminergic innervation is the richest. Such sampling is provided from pooled microdiscs punched out of 8 serial 500 micrometer thick sections of the rat brain. With these conditions, the density of receptor sites (Bmax) was found to be twice that of the whole striatum, without modification of the apparent dissociation constant (Kd) and of the Hills slope. Such a procedure applied to rats with complete 6-hydroxydopamine-induced unilateral nigrostriatal lesions showed a moderate decrease in Bmax in the lesioned side up to 6 days after surgery, whereafter the value of Bmax increased progressively up to the thirtieth day, being then 160% over the control value. Conversely, the apparent Kd decreased significantly from the second to the sixth day postsurgery in the lesioned side, and then increased moderately up to the tenth day and drastically from the twenty-first to the thirtieth day. No change was observed in the corresponding intact side. The modifications appeared chronologically compatible with those corresponding to the behavioral denervation supersensitivity, evidencing drastic binding changes as compared with the whole striatum. The unexpected variations in Kd observed were well correlated with those in Bmax, suggesting that the new available binding sites might be of lower affinity. In the light of all these results, a hypothetical model is proposed.

Changes of the striatal 3H-spiperone binding 3–6 weeks after nigrostriatal denervation and after two years

A complete unilateral lesion of the nigrostriatal pathway by 6-hydroxydopamine injection in the substantia nigra induced a drastic increase in striatal dopaminergic binding sites labelled by 3H-spiperone, 30 days after the lesion. This increase (75% over controls) was time restricted: it was only 39% and 34% over control values at respectively 25 and 35 days after the lesion. Furthermore, 45 days after the destruction of the substantia nigra, the density of labelled sites returned close to the homolateral control values, but remained higher than the contralateral ones, according to the right-left difference found in control animals. Quite later (2 years after the lesion), there was a decrease in the density of labelled sites as compared to the respective homolateral control levels. However, such binding sites tend to remain higher in the striatum of the lesioned side than in the striatum of the intact one, although such a difference was not statistically significant, being very close to the right-left asymmetry observed in control animals. Contrary to our previous results with 3H-Haloperidol, the apparent dissociation constant did not vary significantly, whatever the considered delay after the lesion. These results are discussed in the light of previous results obtained by others and by us.

Cardiovascular effects in the rat of intrathecal injections of apomorphine at the thoracic spinal cord level.

Intrathecal (i.t.) administration of apomorphine at the upper thoracic level lowered blood pressure and heart rate in awake rats. This decrease was dose-dependent and competitively antagonized by haloperidol (i.v. and i.t.) or domperidone (i.t.) but not by domperidone (i.v.). Furthermore, these effects of apomorphine were not affected by alpha- and beta-blocking drugs (i.t.). The results suggest a spinal site, at least in part, for the cardiovascular effect of apomorphine.

Time course of the changes in striatal acetylcholine levels induced by pergolide and haloperidol after lesion of the nigrostriatal dopaminergic pathways in the rat

Lesion of the nigrostriatal dopaminergic pathway in the rat by 6-hydroxydopamine enhances the ability of pergolide to increase striatal acetylcholine levels and prevents the haloperidol-induced decrease in acetylcholine concentrations. This supersensitive response of striatal cholinergic cells is already maximal 6 days after lesion but tends to decrease thereafter. As the time course of the development of the supersensitivity of cholinergic cells differs from that of increased dopamine binding site density, the two are probably not causally related, the former reflecting rather a change occurring beyond the dopamine recognition site.

Cortical ablation fails to influence striatal dopamine target cell supersensitivity induced by nigrostriatal denervation in the rat

The possible influence of the corticostriatal (glutamatergic) pathway on denervation-induced striatal dopamine target cell supersensitivity has been investigated in the rat by measuring the changes in striatal acetylcholine levels induced by the dopamine agonist pergolide and the basal dopamine D2-receptor density after combined 6-hydroxydopamine-induced lesion of the substantia nigra and cortical ablation. Lesion of the nigrostriatal dopaminergic pathway alone enhanced the ability of pergolide (0.06-1 mg/kg i.p.) to increase acetylcholine levels and increased the maximal density of [3H]spiperone binding sites in the striatum. Similar changes in these biochemical parameters were observed after combined cortical ablation and nigral lesion. Cortical ablation by itself slightly diminished acetylcholine levels and reduced by 30% [3H]spiperone binding site density in the striatum. These results indicate that the corticostriatal tract does not influence striatal dopamine target cell supersensitivity caused by dopaminergic denervation.

Long-term effects of nigro-striatal denervation on striatal [3H]haloperidol binding

Abstract [3H]Haloperidol binding measured on pooled microdiscs punched from the rostral and dorsal corpus striatum of rats with unilateral, complete 6-hydroxydopamine-induced nigro-striatal destruction allowed sensitive measurement of the time-dependent fluctuations of the binding patterns related to denervation supersensitivity. Both number of binding sites (Bmax) and apparent dissociation constant (KD) increased 30 days after lesioning. On the other hand, Bmax and KD returned to control levels at 45 days while at later periods only Bmax increased.