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. 2018 Dec;96(12):1862-1875.
doi: 10.1002/jnr.24320. Epub 2018 Aug 31.

Pathophysiology in the Suprachiasmatic Nucleus in Mouse Models of Huntington's Disease

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Free PMC article

Pathophysiology in the Suprachiasmatic Nucleus in Mouse Models of Huntington's Disease

Dika Kuljis et al. J Neurosci Res. .
Free PMC article

Abstract

Disturbances in sleep/wake cycle are a common complaint of individuals with Huntington's disease (HD) and are displayed by HD mouse models. The underlying mechanisms, including the possible role of the circadian timing system, are not well established. The BACHD mouse model of HD exhibits disrupted behavioral and physiological rhythms, including decreased electrical activity in the central circadian clock (suprachiasmatic nucleus, SCN). In this study, electrophysiological techniques were used to explore the ionic underpinning of the reduced spontaneous neural activity in male mice. We found that SCN neural activity rhythms were lost early in the disease progression and was accompanied by loss of the normal daily variation in resting membrane potential in the mutant SCN neurons. The low neural activity could be transiently reversed by direct current injection or application of exogenous N-methyl-d-aspartate (NMDA) thus demonstrating that the neurons have the capacity to discharge at WT levels. Exploring the potassium currents known to regulate the electrical activity of SCN neurons, our most striking finding was that these cells in the mutants exhibited an enhancement in the large-conductance calcium activated K+ (BK) currents. The expression of the pore forming subunit (Kcnma1) of the BK channel was higher in the mutant SCN. We found a similar decrease in daytime electrical activity and enhancement in the magnitude of the BK currents early in disease in another HD mouse model (Q175). These findings suggest that SCN neurons of both HD models exhibit early pathophysiology and that dysregulation of BK current may be responsible.

Keywords: BACHD; BK current; Huntington’s disease; Q175; circadian rhythms; suprachiasmatic nucleus.

Conflict of interest statement

Conflict of Interest Statement:

The authors have no conflict of interest to declare.

Figures

Fig. 1:
Fig. 1:
Depressed daytime spontaneous firing rates (SFR) in BACHD dSCN is not GABA-mediated. (A) Representative traces of dSCN neuron electrical activity recorded during the day (left) or night (right) from young WT (top) and BACHD (bottom) mouse brain slices. (B) Representative trace recorded in presence of the GABAA-R antagonist gabazine (10 μM) with the same organization as (A). (C) Summary of WT (white) and BACHD (grey) dSCN neuron SFR with or without GABAzine. Data are shown as the means ± SEM for each group (bars), the scatter plots show the individual values: WT day (white circles, n = 16), WT night (black circles, n = 14), BACHD day (white triangles, n = 21), BACHD night (black triangles, n = 17). 2-way ANOVA with genotype and time as factors. * = significant difference between genotypes; # = significant difference between day and night.
Fig. 2:
Fig. 2:
BACHD dSCN neurons lose day/night rhythms in resting membrane potential (RMP) and input resistance. (A) Resting Vm of WT (white bars) and BACHD (gray bars) cells recorded in TTX and gabazine. Bar is means ± SEM, individual neuron values overlaid. WT day (white circles, n = 48), WT night (black circles, n = 21), BACHD day (white triangles, n = 33), BACHD night (black triangles, n = 27). (B) Current-voltage response curve for negative current injection steps. WT day (white circles, n = 36), WT night (black circles, n = 26), BACHD day (white triangles, n = 29), and BACHD night (grey triangles, n = 28). (C) Representative trace examples of dSCN neurons responses to negative current injection during the day (left) and night (right). The insert shows the current injection protocol used to evoked the voltage responses. The magnitude of post-inhibitory rebound potentials did not vary between the genotypes. Two-way ANOVA with genotype and time as factors. * = significant difference between day and night; # = significant difference between genotype.
Fig. 3:
Fig. 3:
Direct current injection or NMDA treatment rescues depressed daytime SFR of BACHD dSCN neurons. (A) Traces illustrating that positive current injection (5 or 10pA) increases the SFR of BACHD dSCN neurons during the day. (B) Summary of SFR and (C) Vm responses to current injection for WT (white bars and circles, n = 7) and BACHD (grey bars and white triangles, n = 12) neurons. (D) Example trace of BACHD dSCN neuron response to NMDA treatment (25 μM, 5 min). (E) Summary of NMDA SFR responses for WT (white circles, n = 11) and BACHD cells (white triangles, day, n =11). Bars are mean ± SEM. Two-way ANOVA with genotype and treatment as factors. Data were analyzed using a two-way ANOVA with genotype and treatment as factors. * = significant difference between genotypes; # = significant difference between treated and control.
Fig. 4:
Fig. 4:
The BK potassium (K+) current is enhanced in BACHD dSCN neurons. (A) FDR, and (B) BK current trace examples recorded from WT (left) and BACHD (right) dSCN neurons during the daytime. Voltage step protocol shown to right. Current-voltage response curve summary for (C) FDR (n = 35), and (D) BK (n = 22) currents measured from WT (circles) and BACHD (grey triangles) neurons. Symbols and error bars represent means ± SEM. Two-way ANOVA was used to identify significant effects of genotype and/or voltage step magnitude on evoked currents (P ± 0.05). * = significant difference between genotypes.
Fig. 5:
Fig. 5:
BACHD SCN displayed increased expression of the alpha BK subunit. Kcnma1 expression was modestly higher in the mutant SCN. Gene expression levels of the BK subunits Kcnma1 (A), Kcnmb2 (B), Kcnmb4 (C) were analyzed in WT and BACHD SCN tissue by QPCR, normalized to the geometric mean of the housekeeping genes: Ppia and Rplp0, and are reported as the mean ± SEM of WT (n=6) and BACHD (n=7). When normalized one to the other, the expression levels of Ppia (WT: 0.8440 ± 0.05567; BACHD: 0.8178 ± 0.02205) or Rplp0 (WT: 1.208 ± 0.07056; BACHD: 1.228 ± 0.03333) did not differ between genotypes. * = significant difference between genotypes, t-tests.
Fig. 6:
Fig. 6:
Key findings with the BACHD model were confirmed in the Q175 mouse model. (A) The daily peak of SFR in SCN neurons is reduced in Het and Hom Q175 mice. Using the current-clamp recording technique in the whole-cell configuration, we measured the SFR in dSCN neurons during the day (ZT 5-7) and night (ZT 17-19). Bar graphs plot the mean and SEM for each of the groups. Scatter plots show values for the individual dSCN neurons: WT (white circles, day, n = 10; black circles, night, n = 23); Het Q175 (white upward triangles, day, n = 21; black upward triangles, night, n = 17); Hom Q175 (white downward triangles, day, n = 22; black downward triangles, night, n = 23). The data was analyzed using two-way ANOVA with genotype and time as factors. The Q175 dSCN neurons did not exhibit the day/night difference in SFR seen in WT neurons. * = significant difference between genotypes; # = significant difference between day and night. (B) Current injection (5pA) increased the firing rate in the SCN in WT and Het Q175 mice (n=14 per genotype), but the increase seen in WT is significantly larger than that measured in the Het Q175. Symbols show mean ± SEM of the firing rate before, during, and after current injection. Two-way ANOVA was used to identify significant effects of genotype and treatment on firing rate. * = significant difference between genotypes; # = significant difference between control and treated. (C) BK currents measured in Het Q175 dSCN neurons (n = 9) were significantly larger than those in WT (n = 6) during the day (ZT 5-7). Symbols and error bars represent means ± SEM. Two-way ANOVA was used to identify significant effects of genotype and/or voltage step magnitude on evoked currents. There were significant effects of genotype on the BK current at the 60 and 80 mV steps. * = significant difference between genotypes.

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