STREX (black bars) and ZERO (open bars) mRNA levels expressed as a percentage of total BK channel transcripts in the respective tissue at each developmental time point. Splice variant expression was analysed in mouse: a) spinal cord, b) midbrain, c) cerebellum, d) pons and e) medulla at embryonic day 13 (E13), 15 (E15), 18 (E18) and postnatal days 7 and 35 (P7 and P35 respectively). All data are Means ± S.E.M, n = 5/tissue region. * p < 0.05, ** p < 0.01, compared to respective splice variant expression at P35, Kruskal-Wallis non-parametric test with post hoc Dunn's test for multiple comparisons.
Structures on the Diencephalon and you will Telencephalon
Inside the thalamus and you can hypothalamus a tiny, however, significant, boost in complete BK station expression was observed out-of E15 to help you P35 (Profile 3a 3b). On the other hand, complete BK station mRNA term enhanced nearly ten-flex between embryonic and postnatal stages in front cortex, rear cortex, hippocampus, olfactory bulb, striatum and entorhinal cortex (Shape 3c–h). In every places looked at, there can be a significant developmental downregulation off STREX version mRNA expression (Contour 5). For the frontal cortex, posterior cortex, hippocampus, olfactory bulb, striatum and you may entorhinal cortex this is of this a significant upregulation regarding Zero version mRNA term (Profile 5). Within the thalamus and you can hypothalamus zero significant alterations in No version mRNA expression try seen ranging from E15 and you will P35 (Profile 5).
Developmental regulation of total BK channel mRNA expression in tissues from the diencephalon and telencephalon. Total BK channel mRNA levels expressed as a percentage of postnatal day 35, in mouse a) thalamus, b) hypothalamus, c) frontal cortex, d) posterior cortex, e) hippocampus, f) olfactory bulb, g) striatum and h) entorhinal cortex at embryonic day 13 (E13), 15 (E15), 18 (E18) and postnatal days 7 and 35 (P7 and P35 respectively). All data are Means ± S.E.M, n = 5/tissue region. * p < 0.05, ** p < 0.01, compared to respective P35 data, Kruskal-Wallis non-parametric test with post hoc Dunn's test for multiple comparisons.
Developmental regulation of STREX and ZERO variant splicing in tissues from the diencephalon and telencephalon. STREX (black bars) and ZERO (open bars) mRNA levels expressed as a percentage of total BK channel transcripts in the respective tissue at each developmental time point. Splice variant expression was analysed in mouse: a) mamba apk hile thalamus, b) hypothalamus, c) frontal cortex, d) posterior cortex, e) hippocampus, f) olfactory bulb, g) striatum and h) entorhinal cortex at embryonic day 13 (E13), 15 (E15), 18 (E18) and postnatal days 7 and 35 (P7 and P35 respectively). All data are Means ± S.E.M, n = 5/tissue region. * p < 0.05, ** p < 0.01, compared to respective splice variant expression at P35, Kruskal-Wallis non-parametric test with post hoc Dunn's test for multiple comparisons.
Discussion
The brand new share out-of BK channels to your regulation out-of CNS function was critically based mostly on cell particular, subcellular localisation, intrinsic BK route kinetic characteristics, calcium- and you will voltage sensitivities, and you will regulation by varied mobile signalling routes. Including assortment in the practical attributes from BK avenues, encrypted of the a single gene, can be generated by multiple mechanisms in addition to expression and heterotetrameric set up off distinctive line of splice versions of one’s pore-developing subunit, organization which have regulatory beta subunits and you may signalling complexes and posttranslational controls. This study implies that during murine advancement an adding grounds in order to the newest feeling out of BK channels into CNS mode could well be compliment of power over option splicing of your BK station pore building subunit.
The robust developmental changes in splice variant mRNA expression we observe in multiple CNS regions strongly supports the hypothesis that BK channel splicing is coordinated in the developing CNS and is of functional relevance. In all CNS regions examined, the expression of the STREX variant was significantly down regulated in the face of increasing total BK mRNA levels. In most tissues, such as spinal cord and olfactory bulb, this was accompanied by an upregulation in ZERO variant expression suggesting that splicing decisions to exclude the STREX insert are coordinated across all regions of the developing murine CNS. However, there are important exceptions to this rule such as the cerebellum. In the cerebellum, both STREX and ZERO variant expression is developmentally down regulated resulting in ZERO and STREX variants representing < 10% of total BK channel transcripts at P35. In the cerebellum, developmental upregulation of total BK channel mRNA must be accompanied by an increased expression of other site C2 splice inserts. A similar situation must also occur in tissues such as pons and medulla in which STREX expression declines with no significant change in proportion of ZERO variants when comparing between E13 and P35. Analysis of the splicing decisions in CNS regions with distinct splicing patterns should provide important insights into the mechanisms controlling splicing at site C2 during development.
