Distribution of Smad mRNA and proteins in the rat brain

Smad proteins are known to transduce the action of TGF-β superfamily proteins including TGF-βs, activins, and bone morphogenetic proteins (BMPs). In this study, we examined the expression of Smad1, -2, -3, -4, -5, and -8 mRNA in the rat brain by means of RT-PCR and in situ hybridization (ISH). In addition, we examined the nuclear accumulation of Smad1, -2, -3, -5, and -8 proteins after intracerebroventricular injection of TGF-β1, activin A, or BMP6 with immunohistochemistry to investigate whether TGF-β, activin, and/or BMP activate Smads in the rat brain. RT-PCR analysis revealed that Smad1, -2, -3, -4, -5, and -8 mRNA was expressed in the brain and that the Smad3 and Smad8 mRNA differed in the expression level between brain regions. For example, there were high levels of expression of Smad3 mRNA in the cerebral cortex, caudate putamen/globus pallidus, and cerebellum, but low levels in the thalamus and midbrain. Expression of Smad8 mRNA was higher in the midbrain, cerebellum, and pons/medulla oblongata in comparison to the olfactory bulb, cerebral cortex, caudate putamen/globus pallidus, hippocampus/dentate gyrus, and thalamus. ISH signals for Smad1 mRNA were widely detected in the brain except for a small number of regions including the olfactory tubercle, posterior region of hypothalamus, and cerebellar nuclei. ISH signals for Smad2 mRNA were abundantly observed in several brain regions including the olfactory bulb, piriform cortex, basal ganglia, cingulate cortex, epithalamus, including the pineal gland and medial habenular nuclei, hypothalamus, inferior colliculi of the midbrain, and some nuclei in the pons, cerebellar cortex, and choroid plexus. ISH signals for Smad3 mRNA were also abundantly observed in several brain regions. Especially strong signals for Smad3 mRNA were observed in the olfactory tubercle, piriform cortex, basal ganglia, dentate gyrus, and cingulate cortex. ISH signals for Smad5 and Smad8 mRNA were restricted to a small number of brain regions, the signal intensity of which was weak. ISH signals for Smad4 mRNA were detected in all regions examined. Intracerebroventricular injection of activin A induced nuclear accumulation of Smad2 and Smad3 immunoreactivity in neurons. In contrast, intracerebroventricular injection of TGF-β1 or BMP6 did not induce nuclear accumulation of the immunoreactivity for any Smad in neurons. These results suggest that activin-Smad signaling plays an important role in brain homeostasis.