The correlation of the subcellular localization of dopamine D(1) and D(2) receptors (DA D(1) R, DA D(2) R) with nicotine addiction has not been studied. We demonstrated the ultrasubcellular organelle localization of DA D(1) and D(2) Rs in the caudate-putamen (CPu) area of rat brain in vivo exposed to nicotine (3 mg/day; oral) and passive cigarette smoking (500 ml each; 3 times/day) for 1, 4, and 12 weeks, respectively. Our results revealed DA D(1) R localization in the presynaptic and postsynaptic dendrites, endocytic vesicles, and secretory granules, and DA D(2) R localization in the presynaptic dendrites and vesicles. DA D(1) R immunogold particles were highly decreased in the secretory granules of CPu, and increased in the postsynaptic area and vesicles after prolonged nicotine and smoking exposures, suggesting the strong influence of long time smoking and nicotine exposures on DA D(1) R subcellular organelle localization. DA D(2) R immunoreactivity was comparatively less changed than that of the DA D(1) R. Western blot analysis also showed the differential expression of DA D(1) and D(2) R proteins upon nicotine and smoking exposures as compared to the untreated controls. Taken together, the results for the first time suggests the execution of addictive behavior of nicotine through modulation of mesolimbic dopaminergic system targeting subcellular organelle of DA D(1) and D(2) Rs in the CPu of adult rat brain that may lead to novel therapeutic approaches related to nicotine's neuropsychological disorders including drug addiction.

To confirm the modulation role of GABA(B) on ethanol' effects, we studied the effects of ethanol on the neuronal intracellular signals, protein kinase A (PKA) and cAMP-response element binding protein (CREB), by using a system where GABA(B1) receptors were specifically knocked down in the in vitro cultivated cortical neurons. The results showed that the PKA alpha subunit was increased with ethanol treatment, and could be further increased by administering baclofen and phaclofen. By contrast, baclofen and/or phaclofen could decrease ethanol's up-regulation effects on PKA alpha subunit expression in primary cultured cortical neurons in which the GABA(B1) receptor was specifically knocked down using GABA(B1) receptor RNA interference. Furthermore, these effects could lead to changes of phospho (p)-CREB expression, which showed the same expression pattern as PKA. Finally, we observed changes of GABA(B1), PKA, and p-CREB distribution within the same neuronal cells. These results showed that the GABA(B) receptors are critical to ethanol's cellular effects, which occur via modulating the PKA and CREB transcription pathway, and may be an underlying cause of ethanol's effects.

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Gamma-aminobutyric acid (GABA)(B) receptors appear to influence developmental events, depending on whether they are found at a synapse or in extrasynaptic areas. Little, if anything, is known as to the cellular and subcellular localization of GABA(B1) and GABA(B2) receptors during early fetal development. We used Western blots, immunohistochemistry, and postembedding immunoelectronmicroscopy to investigate fetal rat brain expression and distribution of these receptor proteins. GABA(B1) is expressed as early as gestational day (GD) 11.5 and 12.5, with immunoreactivity found in the all neuroepithelium, and a high expression in the mantel zone and the cortical area's plate; no immunolabeling for GABA(B2) receptor was observed. Our immunogold studies define a pattern of early GABA(B1) receptor protein in dendrite processes, endoplasmic reticulum, and axon terminals of the cortical neuroepithelium on GD 11.5. On GD 12.5, GABA(B1) receptor immunogold was found in dendrite processes, spines and tree, axon terminals, mitochondria, and intracellular organelles of the cortical neuroepithelium. No synapse formation was apparent as no synaptophysin could be found on either GD 11.5 or 12.5. We suggest that GABA(B1) has a functional role in the early fetal brain during neuronal proliferation and migration, and that it is different from the established functional GABA(B) receptor.