In Vivo Electroretinographic (ERG) Studies of the Role of Gabac Receptors in Visual Signal Processing in Mouse Retina, and the Photopic Negative Response of Erg in Humans with Multiple Sclerosis



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Purpose: Experiment 1&2: GABA is a major inhibitory neurotransmitter in central neural system (CNS). Three classes of receptors, GABAA, GABAB and GABAC are all expressed in the retina. This study investigated the role of GABA receptors (GABAR), especially GABACR at different stages of retinal signal processing by studying effects on mouse electroretinogram (ERG) of genetic deletion of GABACRs or using receptor antagonists and agonists. Experiment 3: Multiple sclerosis (MS) is an autoimmune disease of CNS characterized by demyelination and axonal degeneration. 30-70% of MS patients develop optic neuritis (ON). This study used the photopic ERG and compared functional and structural status of inner retina in MS patients.

Methods: Experiments 1&2: Dark-adapted (DA) and light-adapted (LA) ganzfeld brief-flash ERGs were recorded from anesthetized C57BL/6 mice before and after intravitreal injection of GABA, GABAAR antagonist (SR95531), GABABR agonist (baclofen), GABACR antagonists TPMPA, 2-AEMP and 3-APMPA and also from GABACR-/- mice. Negative (n) and positive (p) scotopic threshold responses (STR) were measured at 200 and 110 ms, respectively. After oscillatory potentials (OPs, 50-300 Hz) were removed, b-waves were measured at peak times, 110 ms in DA-ERG, and 50 ms, LA-ERG. OPs' amplitudes were quantified as root mean square (RMS) of extracted OPs. Experiment 3: Photopic ERGs were recorded from 51 MS patients and 33 control subjects. In patients, Humphrey visual fields (HVF) were measured, and peripapillary retinal nerve fiber layer (RNFL) thickness was assessed by optical coherence tomography (OCT) and scanning laser polarimetry (GDx). MS eyes were divided into ON>6 months (n=25), ON<6 months (n=29) and no-ON eye (n=33) groups based on history of ON and time since last ON episode. Thirteen MS patients (13 eyes, ON<6) were re-evaluated one year after the first visit.

Results: Experiments 1&2: GABA injection eliminated STRs and increased b-wave peak amplitude in WT mice, but reduced DA-ERG b-wave amplitudes in GABACR-/- mice. In GABACR-/- mice, or after injection of TPMPA, 2-AEMP or 3-APMPA in WT mice, b-wave maximum amplitudes were attenuated in both DA-ERG and LA-ERG by 30–60%; a-waves were unaffected. OPs’ amplitudes were increased in DA-ERG of GABACR-/- mice by 50%, and in WT mice after TPMPA (~40%) or 2-AEMP (~30%) injection, and 15–35% in LA-ERG. 2-AEMP did not alter nSTR. However, nSTR was enhanced by TPMPA (p=0.04) and 3-APMPA (p=0.04), both partial agonists of GABAB receptors. Baclofen enhanced nSTR (p<0.01) and LA-ERG PhNR (p<0.05). After blockade of GABAARs, an nSTR-like potential was enlarged. TTX partly suppressed the enhanced nSTRs and PhNRs, relating them to spiking activity of inner retinal neurons.

Experiment 3: PhNR amplitudes were lower in ON>6, ON<6 and no-ON eyes (mean±SD, 17.3±7.6, 16.0±6.5, 23.8±9.3μV), than in control eyes (29.8±6.5μV, p<0.001) for a standard stimulus of 1.42 cd-s/m2; a- and b-wave amplitudes were unaffected. PhNR amplitudes were correlated (p<0.05) with HVF Mean Deviation (MD) in ON>6 (r2=0.43) and no-ON eyes (r2=0.10), with similar results for weaker stimuli. PhNR amplitudes were correlated with RNFLT in ON>6 eyes: OCT (r2=0.52, p<0.0001); GDx (r2=0.51, p<0.01), and no-ON eyes: OCT (r2=0.21, p<0.01); GDx (r2=0.17, p<0.05). ON<6 amplitudes were not significantly correlated with other measures, but increased after one year by 5.1±3.1μV (p<0.001), HVF MD by 1.8±2.3 dB (p<0.05), whereas RNFL loss persisted.

Conclusions: Experiment 1&2: GABACRs in On bipolar cells that generate ERG b-waves must be functional for normal maximum b-wave amplitudes to occur. Antagonist effects of newly developed 2-AEMP are similar to those of other antagonists, but with less GABAB agonist properties. Experiment 3: PhNR amplitude in MS patients is significantly reduced in eyes with and without a history of ON. Recovery of function by retinal ganglion cells after acute ON may contribute to visual function improvements in MS patients.



ERG, PhNR, GABA, Retina, Multiple Sclerosis