Michael S. Lidow

Distribution of dopaminergic receptors in the primate cerebral cortex : quantitative autoradiographic analysis using [3H]raclopride, [3H]spiperone and [3H]SCH23390

A widespread distribution of dopamine D1 receptors in the neocortex is well recognized. However, the presence of dopamine D2 receptors in this structure has only recently been established [Martres et al. (1985) Eur. J. Pharmac. 118, 211-219; Lidow et al. (1989) Proc. natn. Acad. Sci. U.S.A. 86, 6412-6416]. In the present paper, a highly specific antagonist, [3H]raclopride, was used for autoradiographic determination of the distribution of D2 receptors in 12 cytoarchitectonic areas of the frontal, parietal, and occipital lobes of the rhesus monkey. A low density of D2-specific [3H]raclopride binding (1.5-4.0 fmol/mg tissue) was detected in all layers of all cortical areas studied. Throughout the entire cortex, the highest density of binding was consistently found in layer V. This is a unique distribution not observed so far for any other neurotransmitter receptor subtype in monkey cerebral cortex, including D1 receptor. In addition, a comparison was made of the distribution of [3H]raclopride and [3H]spiperone, which has been commonly used in previous attempts to label cortical D2 receptors. We found marked differences in the distribution of these two radioligands. In the prefrontal cortex, the pattern of [3H]spiperone binding in the presence of ketanserin resembled the combined distribution of 5-HT1C serotoninergic and alpha 2-adrenergic sites as well as D2 receptors. Thus, [3H]raclopride provides a better estimation of the D2 receptor distribution than does [3H]spiperone. The distribution of D2-specific binding of [3H]raclopride was also compared with the D1-specific binding of [3H]SCH23390 in the presence of mianserin to block labeling to 5-HT2 and 5-HT1C sites. The density of D1-specific [3H]SCH23390 binding was 10-20 times higher than that of D2-specific [3H]raclopride binding throughout the cortex. The densities of both [3H]raclopride and [3H]SCH23390 binding sites display a rostral-caudal gradient with the highest concentrations in prefrontal and the lowest concentrations in the occipital cortex. However, the binding sites of these two ligands had different laminar distributions in all areas examined. In contrast to preferential [3H]raclopride binding in layer V, a bilaminar pattern of [3H]SCH23390 labeling was observed in most cytoarchitectonic areas, with the highest concentrations in supragranular layers I, II and IIIa and infragranular layers V and VI. Whereas [3H]raclopride binding was similar in all cytoarchitectonic areas, [3H]SCH23390 exhibited some region-specific variations in the primary visual and motor cortex. The different regional and laminar distributions of D1 and D2 dopaminergic receptors indicates that they may subserve different aspects of dopamine function in the cerebral cortex.

Layer V neurons bear the majority of mRNAs encoding the five distinct dopamine receptor subtypes in the primate prefrontal cortex

In situ hybridization histochemistry was used to determine the laminar distribution of D1, D2, D3, D4, and D5 dopamine receptor mRNAs in the primate prefrontal cortex and to compare striatal and cortical levels of these messages within the same tissue sections. All five subtypes of dopamine receptor mRNA are present in both the monkey striatum and the cerebral cortex but in different proportions within each structure. Thus, levels of D1 and D2 mRNAs are noticeably stronger in the striatum than in the cortex, whereas D4 and D5 expression is clearly higher in the cortex. The D3 transcripts appear nearly equivalent in the striatum and the cortex. A major finding is that, within the prefrontal cortex, mRNAs encoding all dopamine receptor subtypes are expressed most strongly in layer V. This laminar pattern of mRNA distribution does not hold in all cortical regions. The relatively high levels of mRNAs encoding known dopamine receptor subtypes in the primate cerebral cortex, including the D4 receptor, underscore the importance of this structure as a target for therapeutic actions of antipsychotic drugs. Further, their prominence in layer V of the prefrontal cortex, which contains the corticostriatal and corticotectal projection neurons, provides a neural basis for dopaminergic regulation of the descending control systems. Synapse 28:10–20, 1998.

Adrenergic and serotonergic receptors in aged monkey neocortex

Autoradiography was employed to compare the distribution and density of adrenergic (alpha 1, alpha 2, and beta) and serotonergic (5-HT1 and 5-HT2) receptors in the neocortex of young adult (3 to 10 years of age) and aged (> 20 years of age) rhesus monkeys. The age-related changes in the density of adrenergic and serotonergic sites were area and layer specific. A decrease in the density of alpha 1 receptors occurred only in the superficial layers of the somatosensory cortex, whereas the density of alpha 2 receptors declined in layer I of the prefrontal cortex and in most layers of the motor and somatosensory regions. The increase in beta receptors was largely confined to the deep layers of the motor and somatosensory areas. The density of 5-HT1 sites decreased in most layers of the somatosensory cortex, while 5-HT2 receptors declined in the deep layers of the motor cortex and middle strata of the visual cortex. Overall, adrenergic and serotonergic receptors were least affected in the prefrontal cortex and most compromised in the motor and somatosensory cortex of aged primates.

Distribution of major neurotransmitter receptors in the motor and somatosensory cortex of the rhesus monkey

The in vitro quantitative autoradiographic technique was used to characterize the distributions of alpha 1, alpha 2, beta 1 and beta 2 adrenergic, D1 and D2 dopaminergic, 5-HT1 and 5-HT2 serotonergic, M1 and M2 cholinergic, GABAA and benzodiazepine receptors in the motor (Brodmanns area 4) and somatosensory (Brodmanns areas 3, 1 and 2) cortex of the adult rhesus monkey. All receptor subtypes studied were present throughout all layers of both areas. In the somatosensory cortex, each receptor had its own laminar distribution. Some subtypes of the same receptor (5-HT1 and 5-HT2; alpha 1 and alpha 2) had complementary distributions while others (beta 1 and beta 2; D1 and D2; M1 and M2) had largely overlapping distributions. In contrast, different receptors had remarkably coincidental distributions in the motor cortex. In this area, they all tended to concentrate in layers I, II and the upper part of layer III. However, such coextensive distribution of many types of neurotransmitter receptors is not observed in motor cortex of rats and humans and therefore may be a distinctive feature of motor cortex in the rhesus monkey. The findings described in this paper indicate that somatosensory and motor areas are distinct in their receptor architecture and that receptor autoradiography provides a useful complement to classical histological techniques in elucidating areal differences in the cortex.

Differential quenching and limits of resolution in autoradiograms of brain tissue labeled with3H-,125I- and14C-compounds

Problems in interpretation of autoradiograms generated by ligand binding in brain tissue may be caused by two types of technical limitations: the differential absorption of 3H-generated emissions within the tissue (differential quenching) and the reduced resolution when 125I and 14C are used as isotopes. In the course of our ongoing receptor binding studies in primate brain, we have examined these methodological problems using neocortex of adult rhesus monkey as an example of a complex multilayered brain structure. We have compared: (1) film images produced by brain sections mounted on 3H- and 14C-labeled plastic; (2) autoradiograms of sections labeled with pairs of similar compounds containing 3H, 125I or 14C; and (3) autoradiograms of normal and defatted brain sections. The results indicate that differential absorption of 3H-generated emissions presents a genuine problem for film autoradiography of neocortex of adult monkey when 3H-compounds are used. Particularly significant attenuations of 3H-generated emissions are associated with sublayer IVb of primary visual cortex (Brodmanns are 17) and layers III (deep strata), V and VI of primary motor cortex (Brodmanns area 4). This study provides the necessary corrections for autoradiographic measurements. We also found a loss of resolution associated with use of 125I and 14C, a result that poses a significant problem for analysis of fine laminar patterns of the neocortex in adult monkeys. The use of isotopes with high energy emissions tends to decrease the variations in optical densities within the autoradiograms of cortical sections. Thus, the variations in optical density of autoradiograms of cortices labeled with 125I- and 14C-compounds may not represent the true distribution of these compounds.

α2A‐adrenergic receptors are expressed by diverse cell types in the fetal primate cerebral wall

The cellular elements of the fetal monkey cerebral wall expressing α2A, the most common subtype of the α2 receptor class, were examined by using nonisotopic in situ hybridization and immunohistochemistry with double‐labeling for cell type‐specific markers. At the three embryonic ages examined, E70, E90, and E120, α2A receptors were expressed throughout the embryonic cerebral wall. In the E70 and E90 fetuses, α2A receptors were observed in most cells of the proliferative zones. Some α2A‐positive cells also expressed a proliferation‐associated antigen, Ki‐67, suggesting that the receptors are present in dividing cells. Furthermore, at E90, α2A receptors were detected on fibers passing between the ventricular and subventricular proliferative zones. At all ages studied, α2A receptors were expressed by migrating neurons in the intermediate zone, characterized by a spindle‐like shape, radial alignment, and close association with radial glia. α2A receptors were also expressed by postmigrational microtubule‐associated protein‐2‐positive neurons of the intermediate and subplate zones and the cortical plate. In the marginal zone, α2A receptors were present in the Cajal‐Retzius neurons. Finally, α2A receptors were seen in the glial fibrillary acidic protein‐positive cells at all ages studied. In addition, dopamine‐β‐hydroxylase immunohistochemistry, employed to determine the potential source of noradrenaline in the embryonic cerebral wall, revealed noradrenergic innervation in the marginal, subplate, and intermediate zones of the monkey occipital lobe as early as E70. Based on our observations and available data on α2A signal transduction pathways, we propose that these receptors are involved in regulating the generation, migration, and maturation of cerebral cortical cells. J. Comp. Neurol. 378:493–507, 1997.

D1- and D2 dopaminergic receptors in the developing cerebral cortex of macaque monkey: A film autoradiographic study

Film autoradiography was used to study the distribution of D1- and D2-dopaminergic receptors in the prefrontal association, somatosensory, primary motor and visual regions in the developing cerebral cortex of macaque monkeys. D1 receptors were labeled with [125I]SCH23982, while D2 sites were visualized with [125I]epidepride. D1- and D2-dopaminergic sites are already present in all cortical areas at embryonic day 73, the earliest age observed in this study. In contrast to the adult cortex, where D1 and D2 receptors have different distributions, during development there are substantial similarities in the laminar patterns of these sites. In particular, both D1 and D2 receptors tend to concentrate in the marginal zone and layer V of the developing cortical plate. The autoradiograms also show a high density of D1-dopaminergic sites in the transient ventricular and subventricular zones, where cortical neurons are generated. Although there is a significant rearrangement of the early laminar patterns, the adult distribution of both dopaminergic receptors in most cortical areas is achieved prenatally, soon after all cortical neurons assume their final positions. An early presence in the cerebral wall, a high density in the proliferative zones and fast maturation of the laminar distribution suggests that dopaminergic receptors may be involved in the regulation of cortical development.

Autoradiographic comparison of D1-specific binding of [3H]SCH39166 and SCH23390 in the primate cerebral cortex

Quantitative autoradiography was used to compare the binding of the novel dopamine D1 receptor antagonist, [3H]SCH39166, with that of the widely used radioligand, [3H]SCH23390 (in the presence of ritanserin), in the primate cerebral cortex. Specific binding of both radioligands, determined using SCH23390 or cis-flupentixol as displacing agents, had very similar densities and distributions throughout the cortex. However, the specific binding of [3H]SCH39166 obtained with SCH39166 as a blank was significantly higher than that obtained using SCH23390 or cis-flupentixol as displacing agents in some layers of motor, somatosensory and occipital cortices. In addition, the non-specific binding of [3H]SCH39166 obtained in the presence of an excess of SCH23390 of cis-flupentixol displayed a complex laminar pattern very different from that of the specific binding. These observations suggest that [3H]SCH39166 may have a high affinity to more than the D1 receptor subtype bound by SCH23390 or cis-flupentixol. Also, these additional sites are likely to be different from 5-HT2 or 5-HT1C receptors since the latter sites were not displaced by 1 microM SCH23390.