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Daytime spikes in dopaminergic activity drive rapid mood-cycling in mice.

Sidor MM, Spencer SM, Dzirasa K, Parekh PK, Tye KM, Warden MR, Arey RN, Enwright JF, Jacobsen JP, Kumar S, Remillard EM, Caron MG, Deisseroth K, McClung CA - Mol. Psychiatry (2015)

Bottom Line: Mood-cycling coincides with abnormal daytime spikes in ventral tegmental area (VTA) dopaminergic activity, tyrosine hydroxylase (TH) levels and dopamine synthesis.To determine the significance of daytime increases in VTA dopamine activity to manic behaviors, we developed a novel optogenetic stimulation paradigm that produces a sustained increase in dopamine neuronal activity and find that this induces a manic-like behavioral state.Finally, we show that CLOCK acts as a negative regulator of TH transcription, revealing a novel molecular mechanism underlying cyclic changes in mood-related behavior.

View Article: PubMed Central - PubMed

Affiliation: Department of Psychiatry, University of Pittsburgh Medical School, Pittsburgh, PA, USA.

ABSTRACT
Disruptions in circadian rhythms and dopaminergic activity are involved in the pathophysiology of bipolar disorder, though their interaction remains unclear. Moreover, a lack of animal models that display spontaneous cycling between mood states has hindered our mechanistic understanding of mood switching. Here, we find that mice with a mutation in the circadian Clock gene (ClockΔ19) exhibit rapid mood-cycling, with a profound manic-like phenotype emerging during the day following a period of euthymia at night. Mood-cycling coincides with abnormal daytime spikes in ventral tegmental area (VTA) dopaminergic activity, tyrosine hydroxylase (TH) levels and dopamine synthesis. To determine the significance of daytime increases in VTA dopamine activity to manic behaviors, we developed a novel optogenetic stimulation paradigm that produces a sustained increase in dopamine neuronal activity and find that this induces a manic-like behavioral state. Time-dependent dampening of TH activity during the day reverses manic-related behaviors in ClockΔ19 mice. Finally, we show that CLOCK acts as a negative regulator of TH transcription, revealing a novel molecular mechanism underlying cyclic changes in mood-related behavior. Taken together, these studies have identified a mechanistic connection between circadian gene disruption and the precipitation of manic episodes in bipolar disorder.

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Related in: MedlinePlus

Time specific alterations in VTA dopaminergic activity in ClockΔ19 mice(a) Two-way ANOVA of firing rate found a significant genotype effect (F1,112=12.67, p=0.0006). Post-hoc tests revealed significant differences in the firing rate of dopaminergic neurons during the first 6 h of the light cycle and the last 6 h of the dark cycle (p<0.05 using student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT (n=8) and ClockΔ19 mice (n=9), respectively). (b) Two-way ANOVA of bursting rate found a significant genotype effect (F1,112=8.6, p=0.004) with post-hoc analyses revealing significant differences in the bursting rate of dopaminergic neurons during the first 6 h of the dark cycle (p<0.05, by student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT and ClockΔ19 mice, respectively). (c) Relative abundance of TH mRNA normalized to the expression of Gapdh. Two-way ANOVA revealed a significant genotype effect (F(1,45)=9.42, p=0.004) with a specific increase in TH expression at ZT 4 in ClockΔ19 mice (p< 0.05, n = 3–5 animals/genotype/time point). Diurnal variation was significant in wild-type (WT) mice (CircWave: F(2,27)=10.63, p=0.0004) but not in ClockΔ19 mutants (p>0.05).(d) A significant main effect of time was found for total TH (F(3,28)=5.34, p=0.005), with an increase in ClockΔ19 mouse TH levels at ZT 9 (p < 0.01). Diurnal variation was statistically significant in mutants (CircWave: F(2,17)=5.15, p=0.02). (e) There was a significant effect of time on phosphorylated TH (ser 40) protein (F(3,32)= 6.50, p=0.002), with ClockΔ19 mice exhibiting a specific increase in THser40 levels at ZT 9 (p<0.05, student’s t-test). Diurnal variation in THser40 was statistically significant in mutants (Circwave: F(2,17)=7.18, p=0.005). (f) No differences in phosphorylated TH (ser31) protein levels were found at any time point measured. Inset depicts average protein levels over 24 h. (e–f) Dopamine synthesis assay.(g) Dopamine synthesis was significantly increased in ClockΔ19 mutant mice as measured by L-Dopa in the nucleus accumbens after NSD-1015 administration during the light phase, at ZT 4 (t9 = 2.546, p=0.03). (h) Dopamine synthesis was unaltered in the dorsal striatum of ClockΔ19 mutants (p>0.05, n=5–8 per group; dark phase = ZT 16). White and dark bars below graph represent daytime and night time measurements, respectively.
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Figure 2: Time specific alterations in VTA dopaminergic activity in ClockΔ19 mice(a) Two-way ANOVA of firing rate found a significant genotype effect (F1,112=12.67, p=0.0006). Post-hoc tests revealed significant differences in the firing rate of dopaminergic neurons during the first 6 h of the light cycle and the last 6 h of the dark cycle (p<0.05 using student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT (n=8) and ClockΔ19 mice (n=9), respectively). (b) Two-way ANOVA of bursting rate found a significant genotype effect (F1,112=8.6, p=0.004) with post-hoc analyses revealing significant differences in the bursting rate of dopaminergic neurons during the first 6 h of the dark cycle (p<0.05, by student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT and ClockΔ19 mice, respectively). (c) Relative abundance of TH mRNA normalized to the expression of Gapdh. Two-way ANOVA revealed a significant genotype effect (F(1,45)=9.42, p=0.004) with a specific increase in TH expression at ZT 4 in ClockΔ19 mice (p< 0.05, n = 3–5 animals/genotype/time point). Diurnal variation was significant in wild-type (WT) mice (CircWave: F(2,27)=10.63, p=0.0004) but not in ClockΔ19 mutants (p>0.05).(d) A significant main effect of time was found for total TH (F(3,28)=5.34, p=0.005), with an increase in ClockΔ19 mouse TH levels at ZT 9 (p < 0.01). Diurnal variation was statistically significant in mutants (CircWave: F(2,17)=5.15, p=0.02). (e) There was a significant effect of time on phosphorylated TH (ser 40) protein (F(3,32)= 6.50, p=0.002), with ClockΔ19 mice exhibiting a specific increase in THser40 levels at ZT 9 (p<0.05, student’s t-test). Diurnal variation in THser40 was statistically significant in mutants (Circwave: F(2,17)=7.18, p=0.005). (f) No differences in phosphorylated TH (ser31) protein levels were found at any time point measured. Inset depicts average protein levels over 24 h. (e–f) Dopamine synthesis assay.(g) Dopamine synthesis was significantly increased in ClockΔ19 mutant mice as measured by L-Dopa in the nucleus accumbens after NSD-1015 administration during the light phase, at ZT 4 (t9 = 2.546, p=0.03). (h) Dopamine synthesis was unaltered in the dorsal striatum of ClockΔ19 mutants (p>0.05, n=5–8 per group; dark phase = ZT 16). White and dark bars below graph represent daytime and night time measurements, respectively.

Mentions: To determine how VTA dopamine cell firing changes over the course of the light-dark cycle in tandem with behavioural cycling, in vivo electrophysiological recordings were performed across the 24-hour cycle. VTA dopamine neurons (WT: n=14/14/16/12 and ClockΔ19 mice: n=14/14/17/12 for the number of dopaminergic neurons analyzed across each of the four time bins) were recorded during REM sleep to ensure locomotor activity would not influence dopaminergic firing (Supplemental Figure 2). In addition, neuronal firing was measured during REM sleep rather than anesthetization, as REM sleep is a normal physiological state21, 22. Consistent with previous reports23, ClockΔ19 mice spent significantly less time in REM sleep during the day compared to WT littermates (ZT 0-6, p<0.05; ZT 6-12, p<0.05) and displayed a highly disrupted diurnal rhythm in overall REM sleep (Supplemental Figure 2). Firing rates were elevated in ClockΔ19 mutants throughout the light-dark cycle, reaching significance at ZT 0-6 (p<0.05; Figure 2a). Bursting rates were also significantly elevated in ClockΔ19 mice with the largest effect at ZT 12-18 (p<0.05; Figure 2b). Since our data suggested that genotypic differences in bursting rate were largely driven by a reduced bursting rate in WT dopaminergic neurons, additional firing pattern analyses were performed. When bursting rates were compared in WT mice between ZT 0-6 versus ZT 12-18, a significant reduction in bursting rate was found during the night (ZT 0-6: 2.3±0.4bursts/10 seconds; ZT 12-18: 1.4±0.3bursts/10 seconds) suggesting a diurnal rhythm in activity. Conversely, no differences in bursting rates were observed in ClockΔ19 mice, indicating a loss of diurnal rhythmicity. When differences in firing rate between day and night were compared, ClockΔ19 mice displayed higher dopamine neuron firing during ZT 0-6 (6.5±1.1Hz) than ZT 12-18 (4.8±0.4Hz), with no differences detected in WT mice (ZT 0-6: 3.4±0.5Hz; ZT 12-18: 3.4±0.4Hz). This data demonstrates a significant daytime elevation in tonic dopaminergic activity and a loss in the diurnal rhythm of dopaminergic cell bursting in ClockΔ19 mice.


Daytime spikes in dopaminergic activity drive rapid mood-cycling in mice.

Sidor MM, Spencer SM, Dzirasa K, Parekh PK, Tye KM, Warden MR, Arey RN, Enwright JF, Jacobsen JP, Kumar S, Remillard EM, Caron MG, Deisseroth K, McClung CA - Mol. Psychiatry (2015)

Time specific alterations in VTA dopaminergic activity in ClockΔ19 mice(a) Two-way ANOVA of firing rate found a significant genotype effect (F1,112=12.67, p=0.0006). Post-hoc tests revealed significant differences in the firing rate of dopaminergic neurons during the first 6 h of the light cycle and the last 6 h of the dark cycle (p<0.05 using student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT (n=8) and ClockΔ19 mice (n=9), respectively). (b) Two-way ANOVA of bursting rate found a significant genotype effect (F1,112=8.6, p=0.004) with post-hoc analyses revealing significant differences in the bursting rate of dopaminergic neurons during the first 6 h of the dark cycle (p<0.05, by student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT and ClockΔ19 mice, respectively). (c) Relative abundance of TH mRNA normalized to the expression of Gapdh. Two-way ANOVA revealed a significant genotype effect (F(1,45)=9.42, p=0.004) with a specific increase in TH expression at ZT 4 in ClockΔ19 mice (p< 0.05, n = 3–5 animals/genotype/time point). Diurnal variation was significant in wild-type (WT) mice (CircWave: F(2,27)=10.63, p=0.0004) but not in ClockΔ19 mutants (p>0.05).(d) A significant main effect of time was found for total TH (F(3,28)=5.34, p=0.005), with an increase in ClockΔ19 mouse TH levels at ZT 9 (p < 0.01). Diurnal variation was statistically significant in mutants (CircWave: F(2,17)=5.15, p=0.02). (e) There was a significant effect of time on phosphorylated TH (ser 40) protein (F(3,32)= 6.50, p=0.002), with ClockΔ19 mice exhibiting a specific increase in THser40 levels at ZT 9 (p<0.05, student’s t-test). Diurnal variation in THser40 was statistically significant in mutants (Circwave: F(2,17)=7.18, p=0.005). (f) No differences in phosphorylated TH (ser31) protein levels were found at any time point measured. Inset depicts average protein levels over 24 h. (e–f) Dopamine synthesis assay.(g) Dopamine synthesis was significantly increased in ClockΔ19 mutant mice as measured by L-Dopa in the nucleus accumbens after NSD-1015 administration during the light phase, at ZT 4 (t9 = 2.546, p=0.03). (h) Dopamine synthesis was unaltered in the dorsal striatum of ClockΔ19 mutants (p>0.05, n=5–8 per group; dark phase = ZT 16). White and dark bars below graph represent daytime and night time measurements, respectively.
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Figure 2: Time specific alterations in VTA dopaminergic activity in ClockΔ19 mice(a) Two-way ANOVA of firing rate found a significant genotype effect (F1,112=12.67, p=0.0006). Post-hoc tests revealed significant differences in the firing rate of dopaminergic neurons during the first 6 h of the light cycle and the last 6 h of the dark cycle (p<0.05 using student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT (n=8) and ClockΔ19 mice (n=9), respectively). (b) Two-way ANOVA of bursting rate found a significant genotype effect (F1,112=8.6, p=0.004) with post-hoc analyses revealing significant differences in the bursting rate of dopaminergic neurons during the first 6 h of the dark cycle (p<0.05, by student’s t-test; n=14/14/16/12 and 14/14/17/12 for the number of dopaminergic neurons analyzed in WT and ClockΔ19 mice, respectively). (c) Relative abundance of TH mRNA normalized to the expression of Gapdh. Two-way ANOVA revealed a significant genotype effect (F(1,45)=9.42, p=0.004) with a specific increase in TH expression at ZT 4 in ClockΔ19 mice (p< 0.05, n = 3–5 animals/genotype/time point). Diurnal variation was significant in wild-type (WT) mice (CircWave: F(2,27)=10.63, p=0.0004) but not in ClockΔ19 mutants (p>0.05).(d) A significant main effect of time was found for total TH (F(3,28)=5.34, p=0.005), with an increase in ClockΔ19 mouse TH levels at ZT 9 (p < 0.01). Diurnal variation was statistically significant in mutants (CircWave: F(2,17)=5.15, p=0.02). (e) There was a significant effect of time on phosphorylated TH (ser 40) protein (F(3,32)= 6.50, p=0.002), with ClockΔ19 mice exhibiting a specific increase in THser40 levels at ZT 9 (p<0.05, student’s t-test). Diurnal variation in THser40 was statistically significant in mutants (Circwave: F(2,17)=7.18, p=0.005). (f) No differences in phosphorylated TH (ser31) protein levels were found at any time point measured. Inset depicts average protein levels over 24 h. (e–f) Dopamine synthesis assay.(g) Dopamine synthesis was significantly increased in ClockΔ19 mutant mice as measured by L-Dopa in the nucleus accumbens after NSD-1015 administration during the light phase, at ZT 4 (t9 = 2.546, p=0.03). (h) Dopamine synthesis was unaltered in the dorsal striatum of ClockΔ19 mutants (p>0.05, n=5–8 per group; dark phase = ZT 16). White and dark bars below graph represent daytime and night time measurements, respectively.
Mentions: To determine how VTA dopamine cell firing changes over the course of the light-dark cycle in tandem with behavioural cycling, in vivo electrophysiological recordings were performed across the 24-hour cycle. VTA dopamine neurons (WT: n=14/14/16/12 and ClockΔ19 mice: n=14/14/17/12 for the number of dopaminergic neurons analyzed across each of the four time bins) were recorded during REM sleep to ensure locomotor activity would not influence dopaminergic firing (Supplemental Figure 2). In addition, neuronal firing was measured during REM sleep rather than anesthetization, as REM sleep is a normal physiological state21, 22. Consistent with previous reports23, ClockΔ19 mice spent significantly less time in REM sleep during the day compared to WT littermates (ZT 0-6, p<0.05; ZT 6-12, p<0.05) and displayed a highly disrupted diurnal rhythm in overall REM sleep (Supplemental Figure 2). Firing rates were elevated in ClockΔ19 mutants throughout the light-dark cycle, reaching significance at ZT 0-6 (p<0.05; Figure 2a). Bursting rates were also significantly elevated in ClockΔ19 mice with the largest effect at ZT 12-18 (p<0.05; Figure 2b). Since our data suggested that genotypic differences in bursting rate were largely driven by a reduced bursting rate in WT dopaminergic neurons, additional firing pattern analyses were performed. When bursting rates were compared in WT mice between ZT 0-6 versus ZT 12-18, a significant reduction in bursting rate was found during the night (ZT 0-6: 2.3±0.4bursts/10 seconds; ZT 12-18: 1.4±0.3bursts/10 seconds) suggesting a diurnal rhythm in activity. Conversely, no differences in bursting rates were observed in ClockΔ19 mice, indicating a loss of diurnal rhythmicity. When differences in firing rate between day and night were compared, ClockΔ19 mice displayed higher dopamine neuron firing during ZT 0-6 (6.5±1.1Hz) than ZT 12-18 (4.8±0.4Hz), with no differences detected in WT mice (ZT 0-6: 3.4±0.5Hz; ZT 12-18: 3.4±0.4Hz). This data demonstrates a significant daytime elevation in tonic dopaminergic activity and a loss in the diurnal rhythm of dopaminergic cell bursting in ClockΔ19 mice.

Bottom Line: Mood-cycling coincides with abnormal daytime spikes in ventral tegmental area (VTA) dopaminergic activity, tyrosine hydroxylase (TH) levels and dopamine synthesis.To determine the significance of daytime increases in VTA dopamine activity to manic behaviors, we developed a novel optogenetic stimulation paradigm that produces a sustained increase in dopamine neuronal activity and find that this induces a manic-like behavioral state.Finally, we show that CLOCK acts as a negative regulator of TH transcription, revealing a novel molecular mechanism underlying cyclic changes in mood-related behavior.

View Article: PubMed Central - PubMed

Affiliation: Department of Psychiatry, University of Pittsburgh Medical School, Pittsburgh, PA, USA.

ABSTRACT
Disruptions in circadian rhythms and dopaminergic activity are involved in the pathophysiology of bipolar disorder, though their interaction remains unclear. Moreover, a lack of animal models that display spontaneous cycling between mood states has hindered our mechanistic understanding of mood switching. Here, we find that mice with a mutation in the circadian Clock gene (ClockΔ19) exhibit rapid mood-cycling, with a profound manic-like phenotype emerging during the day following a period of euthymia at night. Mood-cycling coincides with abnormal daytime spikes in ventral tegmental area (VTA) dopaminergic activity, tyrosine hydroxylase (TH) levels and dopamine synthesis. To determine the significance of daytime increases in VTA dopamine activity to manic behaviors, we developed a novel optogenetic stimulation paradigm that produces a sustained increase in dopamine neuronal activity and find that this induces a manic-like behavioral state. Time-dependent dampening of TH activity during the day reverses manic-related behaviors in ClockΔ19 mice. Finally, we show that CLOCK acts as a negative regulator of TH transcription, revealing a novel molecular mechanism underlying cyclic changes in mood-related behavior. Taken together, these studies have identified a mechanistic connection between circadian gene disruption and the precipitation of manic episodes in bipolar disorder.

Show MeSH
Related in: MedlinePlus