Research reportDifferential expression of thymosins β4 and β10 during rat cerebellum postnatal development
Introduction
The β-thymosins are a family of highly conserved small actin-binding peptides [30] widely distributed throughout the vertebrate phylum [14], being also present in some marine invertebrates [28]. Thymosin β4 (Tβ4) and thymosin β10 (Tβ10), the most abundant β-thymosins in mammalian species [37], coexist in some tissues at varying ratios [37] but, in general, Tβ10 is less abundant than Tβ4. Although both peptides share a high degree of sequence homology, they show distinct patterns of expression in several tissues [22], [23], [30].
The presence of both β-thymosins in the mammalian nervous system, during pre- and postnatal development, is now well established. Analysis with specific RNA and protein probes demonstrated that Tβ4 and Tβ10 are expressed in neural tissues from early stages of mammalian embryogenesis [5], [6], [10], [17], [20], [22], [24], [32]. After birth, the levels of Tβ10 mRNA show a more pronounced descent than those of Tβ4 mRNA [19], [22]. In the adult rat forebrain, Tβ4 gene is highly expressed in several areas of the brain including the cerebral cortex, the hippocampal–entorhinal region, the infundibular region, the substantia nigra pars compacta the supraoptic, medial amygdaloid and dorsal premammillary nuclei [7], [16], and in the cerebellum [3]. Moreover, Tβ4 mRNA is induced in some neurons and glial cells following focal brain ischemia [33] and treatment with kainic acid [7]. Tβ10 is also localized in several areas of adult forebrain including hippocampus, neocortex and several brain nuclei [8] and it appears to be absent in the adult cerebellum [3]. Furthermore, treatment of adult rats with kainic acid led to an activation of the Tβ10 gene [8]. Altogether, these results provide strong evidence that β-thymosins play an important role in the control of actin polymerization in both neurons and glial cells. Consequently, these peptides could be involved in processes such as motility, synaptogenesis, axon growth or plastic changes in dendritic spines as well as in the restoration of neuronal circuits and glial activation that occur after brain damage.
The cerebellum is a good model for studying the participation of β-thymosins in processes of neuronal and glial migration and differentiation [1]. The aim of the present study is to increase our knowledge about the spatial and temporal distribution of β-thymosins in the developing and mature rat cerebellum, within an age interval ranging from 7 days after birth to 14 months old, using in situ hybridization and immunohistochemical techniques. Our results show that the temporal and cellular distribution of both β-thymosins are clearly different during postnatal cerebellum development.
Section snippets
Preparation of cerebella sections
All experiments were conducted following the European Union guidelines on animal care and experimentation. Cerebella were obtained from Sprague–Dawley rats reared in the stabularium of the University of Santiago. For immunohistochemistry, we used female rats ranging in age from 7- and 18-day-old postnatal (P7 and P18) to 1-, 4- (adult) and 14-month-old (old). The number of animals of each age utilized in this study was six. Animals were deeply anesthetized with ethyl ether and transcardially
Results
We began our study analyzing the distribution of Tβ4 and Tβ10 mRNAs in the cerebella of adult (2-month-old) and old (1-year-old) rats by in situ hybridization. While the signal in old animals was practically absent, the in situ hybridization analysis revealed a marked difference in the expression pattern of both β-thymosin mRNAs in adult rats (Fig. 1). Thus, the hybridization with the Tβ4 riboprobe demonstrated rather high levels of this transcript in the granular and Purkinje cell layers, and
Discussion
The expression of β-thymosins during rat cerebellar development has been studied by Border et al. [3]. They reported that early in postnatal development (P7 and P14) the expression of Tβ4 and Tβ10 is the highest in the premigratory granule cells of the external granular layer, and that Tβ4 is present in a subset of glia throughout postnatal development, and its synthesis is also induced in maturing Golgi epithelial cells, whereas Tβ10 is only detected at very low levels in a small subpopulation
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