Developmental and Regional Patterns of Expression of Methyl-CpG-Binding Protein (MeCP2) mRNA and Protein in Rodent Brainstem
K.A.Warren; I.C.Solomon.
Abstract
Rett Syndrome (RTT) is a progressive neurodevelopmental disorder caused by mutations in the MECP2 gene. A common symptom observed in RTT patients is breathing dysfunction while awake, which is characterized by intermittent episodes of hyperventilation and central apnea although other respiratory abnormalities (e.g., apneusis, air swallowing) have also been reported. The brainstem (BS) sites responsible for the breathing dysfunction observed in RTT are unknown. Here, we evaluated the developmental and regional expression levels of MECP2 mRNA and protein in BS, including respiratory-related regions. Tissue samples of cortex (control), whole BS, and microdissected BS regions containing the dorsomedial pons (DMP), ventrolateral medulla (VLM), dorsomedial medulla (DMM), and medullary raphe were obtained from neonatal (P0-P20) and adult C57BL/6 mice and/or Sprague-Dawley rats, and processed for total RNA or total homogenate protein. Semi-quantitative RT-PCR and immunoblot analyses of neonatal BS samples from mice demonstrated that MECP2 mRNA and protein were both expressed at each postnatal age; however, the expression levels at P0 were markedly lower than those observed at other developmental stages. Immunoblot analyses further revealed that MECP2 protein expression was substantially higher in cortex than in BS of mice at all ages examined except P0, where no cortical expression was detected (as previously observed). Immunoblot analyses of microdissected BS regions from adult mice and neonatal rat demonstrated MECP2 protein expression in each region for each age evaluated, and further demonstrated regional differences in MECP2 expression with the DMM exhibiting the highest levels, followed by the raphe, and then the DMP and VLM. Our results demonstrate that MECP2 is expressed in BS, including respiratory-related BS regions. We suggest that these developmental and regional expression patterns in the respiratory BS may provide initial insight into the breathing dysfunction in RTT.
Lay Summary
A common symptom observed in RTT girls is altered breathing patterns while awake, with episodes of hyperventilation and stoppage of breathing. The brainstem is the region of the brain which is responsible for the breathing dysfunction observed in RTT. However, the developmental and regional expression levels of MeCP2 remains uninvestigated in these respiratory-related regions. Using tissue samples from young and adult mice, MECP2 mRNA and protein expression in various brain stem regions for each age were evaluated. Ultimately, these findings will provide immediate insight into the breathing dysfunctions found in RTT.