Autoradiographic localization of dopamine uptake sites in the rat brain with 3H-GBR 12935.

The regional distribution of dopamine (DA) uptake sites in the rat brain has been studied by quantitative autoradiography using [3H]GBR 12935 as a ligand. The binding of [3H]GBR 12935 to striatal sections was saturable and of high affinity (Kd = 1.6 nM); it occurred at a single population of sites and possessed the pharmacological features of the DA uptake sites. The highest densities of [3H]GBR 12935 binding sites were found in the caudate-putamen, nucleus accumbens, olfactory tubercle, ventral tegmental area and substantia nigra (especially in the pars compacta). Moderate levels of [3H]GBR 12935 binding were observed in globus pallidus, thalamus, hypothalamus, hippocampus, amygdala (basolateral nucleus) and prefrontal and singular cortices. This regional distribution of [3H]GBR 12935 binding closely correlated with the reported distribution of dopaminergic nerve terminals. The topographical distribution of [3H]GBR 12935 has also been studied in detail in striatal subregions and this distribution was compared, using quantitative TH immunoreactivity, to the density of striatal dopaminergic nerve terminals. There is good overlapping between these two regional distributions, the highest density of both markers was found in the lateral part of the striatum and a similar rostro-caudal gradient has been observed. A dopaminergic denervation caused a complete loss of [3H]GBR 12935 in basal ganglia ipsilateral to the lesion.

Quantitative image analysis with densitometry for immunohistochemistry and autoradiography of receptor binding sites--methodological considerations.

Major technical progress in the development of computer-based image analysis has made possible the entry of autoradiography and immunohistochemistry into a new era where quantification by densitometry has become easily accessible. Autoradiography could become quantitative and displayed adequate reproducibility with the help of emulsion-coated films and the use of scales of standards of known radioactivity exposed and analyzed in parallel to the tissue sections. Immunohistochemistry after revelation by a color-based enzymatic technique can also become quantitative, providing that standardization of the crucial steps of the procedure and calibration through a parallel treatment of a scale of antigen standards can be ensured. Such an approach is described here in the rat with reference to tyrosine hydroxylase (TH), the main synthesizing enzyme for catecholamines, and with dopamine (DA) itself, a catecholaminergic neurotransmitter. The different parts of the procedure, which can influence the results, such as the fixation of the animals by perfusion and the evaluation of the fluctuations via the calibration curve, are discussed in detail. Biological validation of the proposed procedure is described by reference to experiments already well documented biochemically, such as the induction effect of reserpine on TH in the rat locus coeruleus and the depleting effect of alpha-methyltyrosine (AMPT), a well-known blocker of TH activity, on rat striatal DA content. Finally the importance of restricting the measurements to the (pseudo)linear portion of the calibration curve is illustrated by the autoradiographic identification of the differential intrastriatal repartition of the dopaminergic D1 and D2 receptor sites, particularly the dual patch-matrix compartments.

Calcitonin gene-related peptide staining intensity is reduced in rat lumbar motoneurons after spinal cord transection: a quantitative immunocytochemical study.

The expression of Calcitonin gene-related peptide (CGRP) has been demonstrated in motoneurons of several species. We have investigated in adult rats the influence of transection of the spinal cord on CGRP immunoreactivity of motoneurons located below the section. Quantitative analysis has been performed with computer-assisted image analysis. As early as 48 h after the section, CGRP immunoreactivity is modified, and the reduction is maximal after one month. Then, both the number of immunoreactive cells and the intensity of staining increase until the 5th month. It is concluded that the expression of CGRP is under the influence of supraspinal afferents to the motoneuron.

Critical review on quantitative autoradiography of D1 and D2 dopaminergic receptors in the striatum of the mammalian brain: differential localization and plastic changes after pharmacological manipulation and dopaminergic input disruption.

Major technical progress in the development of computer-based image analysis systems has made possible the entry of autoradiographic and immunohistochemical techniques into a new era where quantification via densitometry and morphometry has become easily accessible. In this context, quantitative biochemical data can be adapted to anatomical and histological resolution. This adaptation is most efficient in the neuroscience fields because of the huge importance of cellular communication via neuronal networks in the nervous system. Therefore, any experimental approach to the brain which considers the brain as a 'black box' appears now as very crude. In fact, subtle heterogeneity in the distribution of biochemical markers can now be demonstrated, as illustrated here by the use of quantitative autoradiography of D1 and D2 dopaminergic receptors in the striatum of the mammalian brain. Also, local adaptive changes resulting from chronic blockade of the dopaminergic input can be detected after repeated treatments with dopaminergic antagonists selective for D1 or D2 receptors or with surgical lesioning of the dopaminergic nigrostriatal pathway. The resulting plastic changes are unevenly distributed throughout the striatal target organ and vary according to the mode of suppressing the dopaminergic flow: direct destruction of the dopaminergic pathway or selective pharmacological manipulation without physical elimination of the dopaminergic cells themselves. All these results are discussed and reviewed in light of the most recent reports in this field.