This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Diabetes has been indirectly linked to alterations in b-catenin and to altered sensory and cognitive function. It is however completely unknown, which facets of diabetes causally drive mis-regulation of either IR or Wnt pathways thereby affecting b-catenin's functions in-vivo in central neurons for sensory or mnemonic processing. To this end, the work planned for April 2011 with the initiation of funding, is (1) to determine how the main clinical features of diabetes affect b-catenin expression and activity in the thalamo-cortical-amygdaloid loop during the formation of conditioned fear memories. These studies will allow for clear associations between molecular disruption and compromised behavior. These experiments will also provide me with brain generated signals that will be analyzed in all three years with information theory to determine the extent to which diabetes, via b-catenin signaling, separately or synergistically with other receptor activities, affects the encoding of sensory information versus memory consolidation. In year one, progress will be sought in parallel for both specific aims. References to work planned for early in year two are included but not detailed:
Aim 1 : Determine the extent to which diabetes impairs sensory processing required for sensory learning. In years one, two and three, untreated diabetic (db/db) mice, treated (db/db) and non-diabetic controls, along with mice used for generating conditional knockouts for b-catenin will be trained and tested in tone-driven fear-conditioning to quantify memory acquisition. Multi-site, multi-electrode neurophysiological recordings will be carried out in awake animals for tone-induced neural responses. In year one, I will focus on the most challenging of the recordings, the medial geniculate nucleus (MGN). For each anatomical site, I will conduct recordings in two treated db/db groups and three conditions of conditional knockout where b-catenin is removed from three separate anatomical locations. Diabetic mice will be managed by the CoBRE core facility either with the insulin sensitizer Thiazolidinedione or will receive systemic insulin supplements. In all years, spectro-temporal receptive fields (STRFs) will be computed to determine how the tuning of neurons may be altered as a result of diabetes or diabetes treatment, relative to controls. All recordings will also be analyzed with linear pattern classifiers and for mutual information to reveal the effectiveness of the neural coding in each of these brain areas, for each condition, during sensory processing (training) and memory consolidation (testing). These studies will elucidate the causal interaction between changes in IR signaling and downstream activities of the Wnt pathway that alter availability of b-catenin. At the beginning of year two, I will complete the complement of recordings from the primary auditory cortex (A1) and the basolateral amygdala (BLA). Also by the beginning of year two, I will initiate experiments with excised floxed b-catenin by transfecting adenovirus expressing Cre recombinase and a green florescent protein cassette (Baylor Vector Development Lab) directly into the MGN, A1 or the BLA. Recordings will be made and processed as described above.
Aim 2 : Determine how diabetes, via Wnt signaling, alters the biophysical properties of sensory-driven neurons. In year one, I will carry out whole-cell patch-clamp recordings in brain slices containing MGN, A1 and BLA from control animals. Early in year two, I will carry out sufficient recordings for preliminary data for the R01 submission from brain slices from db/db mice and animals bearing a floxed b-catenin construct. Slices will be obtained from animals that have undergone acute versus prolonged states of altered insulin availability, hypo or hyperglycemia.
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