Tinnitus is associated with increased extracellular matrix density in the auditory cortex of Mongolian gerbils

Abstract

Most scientists agree that subjective tinnitus is the pathological result of an interaction of damage to the peripheral auditory system and central neuroplastic adaptations. Here we investigate such tinnitus related adaptations in the primary auditory cortex (AC) 7 and 13 days after noise trauma induction of tinnitus by quantifying the density of the extracellular matrix (ECM) in the AC of Mongolian gerbils (Meriones unguiculatus). The ECM density has been shown to be relevant for neuroplastic processes and synaptic stability within the cortex. We utilized a mild monaural acoustic noise trauma in overall 22 gerbils to induce tinnitus and a sham exposure in 16 control (C) animals. Tinnitus was assessed by a behavioral response paradigm. Animals were separated for a presence (T) or absence (NT) of a tinnitus percept by a behavioral task. The ECM density 7 and 13 days after trauma was quantified using immunofluorescence luminance of Wisteria floribunda lectin-fluoresceine-5-isothiocyanate (WFA-FITC) on histological slices of the primary AC, relative to the non-auditory brainstem as a reference area. At both timepoints, we found that the WFA-FITC luminance of the AC of NT animals was not significantly different from that of C animals. However, we found a significant increase of luminance in T animals' ACs compared to NT or C animals' cortices. This effect was found exclusively on the AC side contralateral to the trauma ear. These results point to a hemisphere specific process of stabilization of synaptic connections in primary AC, which may be involved in the chronic manifestation of tinnitus.

Keywords: Animal model; Auditory cortex; Extracellular matrix; Rodents; Tinnitus.

Fig. 1

Exemplary WFA-FITC marked brain slices from a control (left panel), non-tinnitus (center panel) and tinnitus animal (right panel) 13 days after the trauma. Rectangles identify the evaluated cortical (turquoise)/brainstem reference (yellow) areas. White scale indicates 1 mm. Note that the luminance of the slices could vary, therefore only the ratio of cortex to brainstem luminance is used for further analyses

Fig. 2

A 7 d after trauma results of 2-factorial ANOVAs of mean hearing loss (HL in dB) of plugged control ears (upper panels) and (sham-) trauma ears (lower panels) with the factors group (NT, T, C; color-coded) and frequency (in kHz). Asterisks above horizontal lines give Tukey post-hoc test values (**p < 0.01) and asterisks next to vertical lines give significant single sample t-test values against 0 (**p < 0.01). B Results of GPIAS tests (effect size in AU) for all animals in the three groups over the five tested frequencies. C Separation of non-significant (NT) and significantly negative (T) effect size values across the five tested frequencies

Fig. 3

A 13 d after trauma results of 2-factorial ANOVAs of mean hearing loss (HL in dB) of plugged control ears (upper panels) and (sham-) trauma ears (lower panels) with the factors group (NT, T, C; color-coded) and frequency (in kHz). Asterisks above horizontal lines give Tukey post-hoc test values (**p < 0.01) and asterisks next to vertical lines give significant single sample t-test values against 0 (*p < 0.05; ***p < 0.001). B Results of GPIAS tests (effect size in AU) for all animals in the three groups over the five tested frequencies. C Separation of non-significant (NT) and significantly negative (T) effect size values across the five tested frequencies

Fig. 4

Interaction plots of the two-factorial ANOVAs of the normal distributed luminance ratio for the factors animal group and cortex side for both observation times separately. A The interaction on day 7 after trauma, the F statistics is given in black. The two colored F-statistics show the results of the Bonferroni corrected one-factorial ANOVAs of ipsi- (blue) and contralateral (red) luminance ratios of the three animal groups. B The interaction on day 13 after trauma, colors as above. Colored asterisks and numbers in both panels indicate the significance levels and tendencies of the Tukey post-hoc tests in the one-factorial ANOVAS: * p < 0.05, ** p < 0.01