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Repeated exposure to cocaine produces an enduring increase in dendritic spine density in adult rat nucleus accumbens. It has been shown previously that chronic cocaine administration increases the expression of cyclin-dependent kinase-5 in this brain region and that this neuronal protein kinase regulates cocaine-induced locomotor activity. Moreover, cyclin-dependent kinase-5 has been implicated in neuronal function and synaptic plasticity. Therefore, we studied the involvement of this enzyme in cocaine's effect on dendritic spine density. Adult male rats, receiving intra-accumbens infusion of the cyclin-dependent kinase-5 inhibitor roscovitine or saline, were administered a 28-day cocaine treatment regimen. Animals were killed 24-48 h after the final cocaine injection and their brains removed and processed for Golgi-Cox impregnation. Our findings demonstrate that roscovitine attenuates cocaine-induced dendritic spine outgrowth in nucleus accumbens core and shell and such inhibition reduces spine density in nucleus accumbens shell of control animals. These data indicate that cyclin-dependent kinase-5 is involved in regulation of, as well as cocaine-induced changes in, dendritic spine density., (Copyright 2003 IBRO)

DARPP-32 (dopamine- and cyclic AMP-regulated phosphoprotein, apparent molecular weight of 32,000) is part of the D1 dopamine receptor signal transduction cascade. Both the D1 receptor and DARPP-32 are found in the caudate putamen, but it is not known if they co-localize in the medium-sized spiny neurons. In the present study, double-labelling immunocytochemistry was used to simultaneously localize the D1 receptor and DARPP-32 in the rat caudate-putamen. The neuropil was heavily and uniformly immunoreactive for both the D1 receptor and DARPP-32. All cell bodies immunopositive for the D1 receptor were immunopositive for DARPP-32. The D1 receptor was not detectable, however, in nearly half of the DARPP-32-containing cell bodies. DARPP-32 is present in striatopallidal and striatonigral projections. The D1 receptor co-localized with DARPP-32 in fibres of the entopeduncular nucleus and the pars reticulata of the substantia nigra. In the globus pallidus, however, D1 receptor immunoreactivity was barely detectable, while DARPP-32 immunolabelling of axons and axon terminals was intense. These data suggest that the striatal somata containing both the D1 receptor and DARPP-32 project to the entopeduncular nucleus and substantia nigra, whereas somata containing only DARPP-32 immunoreactivity project to the globus pallidus. Thus, the differences in expression of the D1 receptor and of DARPP-32 within striatal cell bodies are likely reflected in their projections. The co-localization of the D1 receptor and DARPP-32 is consistent with the known regulation of DARPP-32 phosphorylation by D1 receptor activation. The demonstration of a large population of striatal neurons that contain DARPP-32 but apparently do not contain D1 receptors substantiates the premise that these cells have an alternative signal transduction pathway. Subsequent studies are needed to search for a signal transduction pathway for these neurons analogous to the dopamine D1 receptor pathway.

The borders of the ventral striatum and ventral pallidum of the rat brain have been studied with immunocytochemistry for two protein markers that are present in these regions. One of these, DARPP-32 (dopamine and cyclic AMP-regulated phosphoprotein, Mr 32,000), is specifically enriched in medium-sized spiny neurons of the neostriatum and in the projection of these neurons upon pallidal regions. The second protein, synaptophysin, a marker for nerve terminals, effectively labels pallidal synapses. In the ventral striatum, neurons strongly immunoreactive for DARPP-32 were densely packed throughout its three components, i.e., the fundus striati and the nucleus accumbens septi, the olfactory tubercle, and the cell bridges that connect the tubercle with the overlying caudatoputamen and the nucleus accumbens. In the ventral pallidum, axons and axon terminals immunoreactive for DARPP-32 and axon terminals immunoreactive for synaptophysin were clearly delineated. As defined by these markers, the ventral pallidum was traced rostroventrally from the globus pallidus to the superficial layers of the olfactory tubercle, medially to the insula Calleja magna and the lateral septum, laterally to the pyriform cortex, and caudally to the anterior amygdaloid area. The ventral striatum and pallidum were densely intermingled in parts of the olfactory tubercle and medial forebrain bundle regions, and clearly separated in more caudal regions. The insulae Callejae did not contain typical striatal or pallidal staining patterns. Our results indicate that the ventral striatopallidal complex in the rat extends both rostrocaudally and dorsoventrally, in a highly complex, intermingled fashion, throughout most of the basal forebrain.

Tyrosine hydroxylase-like, dopamine- and cyclic AMP-regulated phosphoprotein (Mr = 32,000)-like and enkephalin-like immunoreactive profiles and their codistribution have been evaluated at three rostrocaudal levels of the rat neostriatum by means of a computer-assisted morphometrical method, which allows an objective definition of high density/intensity patches using specific antibodies in combination with the peroxidase-antiperoxidase technique. Our results show that both tyrosine hydroxylase-like, dopamine- and cyclic AMP-regulated phosphoprotein-like and enkephalin-like profiles are organized in patches in the rat neostriatum. In the marginal zone, the tyrosine hydroxylase-like immunoreactive and dopamine- and cyclic AMP-regulated phosphoprotein-like immunoreactive patches both occupied a large part of the total area. Moreover, in this zone, these putative markers for pre- and postsynaptic elements of dopaminergic synapses also showed a complete spatial overlap. In contrast, the enkephalin-like immunoreactive patches in the marginal zone occupied a smaller area, and showed only an incomplete, albeit significant overlap with the tyrosine hydroxylase-like immunoreactive/dopamine- and cyclic AMP-regulated phosphoprotein-like immunoreactive system. In the central zone, tyrosine hydroxylase-like immunoreactive, dopamine- and cyclic AMP-regulated phosphoprotein-like immunoreactive and enkephalin-like immunoreactive patches occupied a much smaller part of the total area than did those in the marginal zone. Within the central zone, enkephalin-like immunoreactive patches occupied a significantly larger area than did the tyrosine hydroxylase-like immunoreactive and dopamine- and cyclic AMP-regulated phosphoprotein-like immunoreactive patches. No consistent pattern of overlap between the three different staining patterns could be seen in the central zone, probably due to the small, inconsistent size of the patches. Trend analysis showed a consistent trend of more tyrosine hydroxylase-like immunoreactive and dopamine- and cyclic AMP-regulated phosphoprotein-like immunoreactive patches in the dorsal than in the ventral striatum, and a trend of more enkephalin-like immunoreactive patches in the rostral than in the caudal striatum. Our data thus demonstrate that, by using computer-assisted morphometrical techniques, it is possible to describe a non-homogenous but overlapping distribution of tyrosine hydroxylase-like immunoreactive and dopamine- and cyclic AMP-regulated phosphoprotein-like immunoreactive patches in the rat neostriatum.(ABSTRACT TRUNCATED AT 400 WORDS)