Dopaminergic Neurons Exhibit an Age-Dependent Decline in Electrophysiological Parameters in the MitoPark Mouse Model of Parkinson's Disease

Pacemaker firing properties are altered in dopaminergic neurons from MitoPark mice. We performed loose cell-attached recordings to monitor the firing properties of dopaminergic neurons in brain slices from MitoPark and control mice. A sample trace (A) from a 15.1-week-old control mouse is shown in contrast to a trace from a 12.4-week-old MitoPark mouse (B) that showed disrupted rhythmicity. Linear regression data indicated an elevation in firing rate with increasing age, with several neurons showing a dramatically higher firing rate in MitoPark mice (C). Firing rates did not significantly differ when data were pooled into age groups (D). Single-cell data of the logarithm of the coefficient of variation of the interspike interval indicated elevated variability in the MitoPark mice that did not change with age (E). When these data were grouped into bins based on mouse age, interspike intervals from MitoPark mice were significantly more variable at 11–15 and 16+ weeks of age (F). The width of the extracellular waveforms was also calculated (G, blue line; sample traces from a 15.1-week-old control and a 15.6 week-old MitoPark mouse), and linear regression revealed an age-dependent decrease in MitoPark mice (H). When grouped into bins, spike widths were significantly decreased in MitoPark mice >16 weeks of age (I). **p < 0.01, ***p < 0.001, ****p < 0.0001.

Slow intrinsic conductances are reduced in dopaminergic neurons from MitoPark mice. The HCN channel-mediated I_h was measured by applying a 1 s hyperpolarizing step from −55 to −105 mV (A). Dopaminergic neurons from MitoPark mice exhibited a globally lower I_h either when analyzed with linear regression (B) or when data were grouped into bins (C). The small conductance SK channel-mediated AHC was measured by applying a 100 ms depolarizing step from −55 to 0 mV (D). Neurons from MitoPark mice exhibited lower area under the curve (charge) that did not depend on age (E, F). *p < 0.05, ***p < 0.001.

D2 dopamine autoreceptor-mediated currents are reduced in dopaminergic neurons from MitoPark mice. D2 receptor-mediated outward currents were measured in dopaminergic neurons from MitoPark and control mice by applying a maximally effective iontophoresis of dopamine. Sample traces from a control (16.4 weeks) and a MitoPark (15 weeks) mouse indicate smaller peak currents in neurons from MitoPark mice (A). Dopamine neurons from MitoPark mice exhibited smaller peak currents across age compared with controls (B), an effect that reached significance at ages >11 weeks (C). ****p < 0.0001.

Amphetamine-induced currents are dramatically reduced in dopaminergic neurons from MitoPark mice. A large concentration (30 μm) of amphetamine was applied to rapidly induce an outward current through dopamine efflux. After the outward current plateaued for several seconds, dopamine was applied by iontophoresis to achieve the maximum possible outward current. A sample trace (A, left) from a 23.6-week-old control mouse indicates how the amphetamine-induced outward current (green) was measured compared with the total D2 receptor-mediated current (blue). For comparison, a sample trace (A, right) is also shown from a 13.4-week-old MitoPark mouse. The amphetamine-induced currents were consistently smaller in recordings from MitoPark mice and decreased with age (B). Amphetamine-induced currents were significantly smaller as early as 6–10 weeks of age, and strikingly were eliminated by 16 weeks of age (C). *p < 0.05, ****p < 0.0001.

Dopaminergic neurons from MitoPark mice exhibit smaller dopamine-mediated IPSCs. IPSCs were electrically evoked using a bipolar stimulating electrode placed caudal to the target dopamine neuron in the presence of GABA, glutamate, and nicotinic acetylcholine receptor blockers. Sample traces (A) are from a 14-week-old control mouse and a 13.4-week-old MitoPark mouse. Stimulus response curves were constructed with increasing stimulus intensities. At all age groups IPSCs from MitoPark mice were smaller as indicated by downward shift in the stimulus response curves (B–D). **p < 0.01, ***p < 0.001, ****p < 0.0001.

Verified tyrosine hydroxylase-positive neurons from MitoPark mice exhibit altered electrophysiological parameters. In a separate experiment, neurons were filled with 0.2% neurobiotin and recorded with an emphasis on obtaining multiple dependent electrophysiological measures from each cell. Cells were later stained (A) for neurobiotin (left) and TH (middle), and cells that definitively co-stained for both markers (right) were analyzed for their electrophysiological parameters. Consistent with previous results, recordings in neurons from MitoPark mice exhibited decrease cell capacitance (B), increased input resistance (C), and decreased I_h (D), spike width (E), and peak current observed in response to iontophoresis of dopamine (F). Two-tailed unpaired t tests. **p < 0.01, ***p < 0.001.