Banca de QUALIFICAÇÃO: INGRID NOGUEIRA SOUSA

Uma banca de QUALIFICAÇÃO de MESTRADO foi cadastrada pelo programa.
DISCENTE : INGRID NOGUEIRA SOUSA
DATA : 01/07/2019
HORA: 09:30
LOCAL: INSTITUTO DO CEREBRO
TÍTULO:

CHARACTERIZATION OF CELL TYPES AFFECTED BY NOISE-INDUCED TINNITUS IN THE AUDITORY CORTEX OF MICE

PALAVRAS-CHAVES:

Tinnitus, Auditory Cortex, Patch Clamp, Pyramidal cells and Martinotti cells

PÁGINAS: 65
GRANDE ÁREA: Ciências Biológicas
ÁREA: Fisiologia
RESUMO:

Tinnitus is an abnormal state of nerve cell activity of the auditory system, leading to perception of phantom sounds, such as ringing of the ears. Although tinnitus perception is not harmful per se, it can lead to severe psychological stress, anxiety and depression. Several studies indicate the auditory cortex as a potential target for transmagnetical stimulation to alleviate tinnitus perception, yet little is known of how tinnitus alters cortical circuits. Here we investigate cellular populations of the primary auditory cortex (A1) in a mouse model of noise-induced tinnitus of adult mice (p38-p52) that does not generate hearing loss. Our goal was to identify specific subtypes of cortical pyramidal cells (PCs) and inhibitory Martinotti cells that are affected by a noise-trauma (4-20kHz, 90dB, 1,5 hrs). To achieve this, we use two strategies: 1) electrophysiological classification of randomly patched layer 5 pyramidal cells in slices from control and a tinnitus-induced group, 2) using an activity-activated calcium integrator (CaMPARI),a fluorescent protein that photoconverts from green to red when stimulated by violet light (~400nm) in the presence of high intracellular calcium concentration. Whole-cell patch clamp recordings from a total of 107 cells (pyramidal cells; control: n=46, tinnitus-group: n=41; Martinotti cells; control: n=8, tinnitus-group: n=12) from layer 5 of A1. Pyramidal cells were characterized based on the size and shape of their soma, location and firing features while Martinotti cells were identified by red fluorescence, using Chrna2-tomato positive animals, a specific marker for this type of cells of A1 layer 5, and also by electrophysiological characteristics. Analyzing firing properties (sag, rebound potential and afterhyperpolarization) allowed us to subdivide pyramidal cells into type A and B pyramidal cells. Type A cells corresponds to cells with large afterhyperpolarizations (AHPs), prominent sags, and pronounced rebound afterdepolarization (ADP). While type B pyramidal cells corresponds to cells with the absence of AHP or ADP, and small hyperpolarization sags. Comparing the two types of pyramidal cells in control slices showed type A PCs (n=11) to have a more depolarized membrane potential compared to type B PCs (A: -65,7±1,5mV, B: -72,3±1,4mV, p=0,003), a lower Rheobase (A: 68,0±4,3pA, B: 108,4±9,5pA, p=0,001), a higher initial firing frequency (A: 64,9±10,9Hz, B: 33,2±3,8Hz, p=0,008) and higher steady state frequency (A: 20,3±1,8Hz, B: 13,3±1,3Hz, p=0,004) compared to type B PCs (n=13). When comparing electrophysiological properties between cells from noise overexposed animals and control animals, we found type A PCs from noise-overexposed (no) animals to have a lower steady state frequency (noA: 16,1±1,2Hz, n=19) than control type A PCs (p=0,050). Type B PCs from noise-overexposed animals showed increase on steady state frequency (noB: 19,5±2,4Hz, n=22) compared to type B PCs from the control animals (p=0,048). After noise-overexposure the difference in resting membrane potential and rheobase between type A and B remained, however the difference in initial and steady state frequency was abolished. Cells from control younger animals (p16-p23) show similar results when compared to the noise-overexposed in a matter of initial (A: 61,9±6,4Hz, B: 46,0±4,15Hz) and steady state frequency (A: 17,9±1,4Hz, B: 17,7±2,3Hz). Interestingly, preliminary data from Martinotti cells from noise-overexposed (noM) animals show a higher initial (noM: 80,5±3,4Hz, M: 70,05±6,5Hz) and steady state frequency (noM: 33,5±4,9Hz, M: 20,35±4,4Hz) in comparison to the control group. Using CaMPARI (n=4 mice) we could not verify specific cell types affected by noise-overexposure, but we could observe cells with higher activity compared to neighboring cells, indicating that noise-overexposure does not affect cells uniformly in layer 5-6 of the A1. Together, these results are a first step in identifying specific cortical neurons affected by acoustic trauma and quantify the electrophysiological differences seen for each subtype. To understand the cellular mechanisms of tinnitus is crucial for improving treatments of tinnitus using cortical stimulation.

MEMBROS DA BANCA:
Interno - 2394627 - EDUARDO BOUTH SEQUERRA
Presidente - 1976236 - EMELIE KATARINA SVAHN LEAO
Interno - 2183828 - TARCISO ANDRE FERREIRA VELHO