The effects of MeCP2 deficiency on the strength of specific microcircuits in mouse motor/frontal cortex

The mammalian neocortex is characterized by a laminar structure, which provides an organization scheme for local synaptic circuits. The first chapter of my thesis research focused on developing a general approach for estimating layer-specific connectivity in cortical circuits and applying it to mouse motor cortex. From these data I computed a laminar presynaptic-postsynaptic connectivity matrix, Wpost,pre, revealing a complement of stereotypic pathways dominated by layer 2 outflow to deeper layers. Comparisons between the major synaptic pathways in motor and somatosensory cortex revealed fundamentally different organization schemes in each, with motor cortex organization dominated by a =top-down' pathway and somatosensory cortex dominated by a 'bottom-up' pathway.;The second and third chapters of my Ph.D. research utilized the cortical circuit connectivity matrix found for motor cortex to assess abnormalities in intracortical synaptic pathways in mouse models of Rett syndrome (RTT). RTT, a rare neurogenetic disorder primarily affecting girls, arises from mutations in the gene encoding methyl-CpG-binding protein 2. Motor and cognitive systems are particularly affected in this disorder, pointing to cortical involvement. In the second part of my thesis I examined synaptic circuits of layer (L) 2/3 pyramidal neurons in the motor-frontal cortical area of MeCP2-null mice (3-4 weeks postnatal age). I mapped local excitatory input to layer L2/3 pyramidal neurons using glutamate uncaging and laser scanning photostimulation, and compared input maps recorded from MeCP2-null and wild type mice. Synaptic input from local excitatory sources was significantly reduced in the mutants, both for horizontal pathways from laterally adjacent L2/3 sites, and for ascending pathways from subjacent L3/5A sites. The reduction in input from L2/3 represented a reduction in synaptic connectivity, while the reduction from L3/5A resulted from reduced presynaptic excitability. These findings provide evidence that MeCP2 deficiency is associated with reduced synaptic transmission in excitatory networks, and demonstrate that cortical circuits in a motor-frontal area are affected in this manner.;The final chapter of my thesis focused on cell-specific affects of MeCP2-deficiency on cortical pathway strength in motor cortex. I used in utero transfection of short hairpin RNA constructs to knock down MeCP2 expression in a sparsely distributed subset of L2/3 pyramidal neurons in wild type mice, and compared input maps recorded from transfected-untransfected pairs of neighboring neurons. The effect of MeCP2 deficiency on local excitatory input pathways was severe, with an average reduction in excitatory synaptic input from middle cortical layers (L3/5A) of greater than 30% compared to MeCP2-replete controls. MeCP2 deficiency primarily affected the strength, rather than the topography, of excitatory intracortical pathways. Inhibitory synaptic inputs and intrinsic eletrophysiological properties were unaffected in the MeCP2-knockdown neurons. These studies indicate that MeCP2 deficiency in individual postsynaptic cortical pyramidal neurons is sufficient to induce a pathological synaptic defect in excitatory intracortical circuits. However, the synaptic pathology is not identical to that found in the MeCP2-null animals, indicating that cell-specific affects of MeCP-deficiency are likely accompanied by global homeostatic responses in cortices which completely lack MeCP2 function.