Photoperiodic effects on monoamine signaling and gene expression throughout development in the serotonin and dopamine systems

Abstract

Photoperiod or the duration of daylight has been implicated as a risk factor in the development of mood disorders. The dopamine and serotonin systems are impacted by photoperiod and are consistently associated with affective disorders. Hence, we evaluated, at multiple stages of postnatal development, the expression of key dopaminergic (TH) and serotonergic (Tph2, SERT, and Pet-1) genes, and midbrain monoamine content in mice raised under control Equinox (LD 12:12), Short winter-like (LD 8:16), or Long summer-like (LD 16:8) photoperiods. Focusing in early adulthood, we evaluated the midbrain levels of these serotonergic genes, and also assayed these gene levels in the dorsal raphe nucleus (DRN) with RNAScope. Mice that developed under Short photoperiods demonstrated elevated midbrain TH expression levels, specifically during perinatal development compared to mice raised under Long photoperiods, and significantly decreased serotonin and dopamine content throughout the course of development. In adulthood, Long photoperiod mice demonstrated decreased midbrain Tph2 and SERT expression levels and reduced Tph2 levels in the DRN compared Short photoperiod mice. Thus, evaluating gene × environment interactions in the dopaminergic and serotonergic systems during multiple stages of development may lead to novel insights into the underlying mechanisms in the development of affective disorders.

Figure 1

Photoperiod paradigm across development. For developmental quantitative RTPCR and midbrain monoamine content experiments, animals developed under either an (A) Equinox LD 12:12, (B) Long LD 16:8, or (C) Short LD 8:16 photoperiod. Mice developed under these photoperiods from embryonic day 0 (E0) and were evaluated at postnatal days (i) 8, (ii) 18, or (iii) 35 (i.e. P8, P18, P35). L = light, D = dark (i.e. LD 16:8 means Light for 16 h, Dark for 8 h).

Figure 2

Perinatal Short photoperiod significantly increased midbrain TH levels. Mice developed under either Equinox (Eq), Long (L), or Short (S) photoperiods and TH expression levels were measured at three time points: P8, P18, and P35. The significant levels were as follows: (****p < 0.0001). Reference gene for relative expression was β-actin.

Figure 3

Short photoperiod reduced midbrain monoamine content at multiple stages of development. (A) 5-HT content, (B) 5-HIAA levels, (C) DA content, and (D) DOPAC levels in the midbrain. Mice developed under either Equinox (Eq), Long (L), or Short (S) photoperiods and monoamine levels were measured at three time points: P8, P18, and P35. The significant levels were as follows: (**p < 0.01, ***p < 0.001). The significance levels in (A) are shown for Long (L) to Equinox (Eq) at P18, and for Long (S) to Short (S) comparisons at P35. In addition, a significant Holm–Sidak’s post hoc test revealed differences between Equinox and Short (p = 0.0014) groups for (A) 5-HT content at P35. The significance levels in (B) are shown for Long (L) to Short (S) comparisons at P18. The significance levels in (C) and (D) are shown for Long (L) to Short (S) comparisons at P35. Lastly, a significant Holm-Sidak’s post hoc test revealed differences between Equinox and Short (p = 0.0493) groups for (C) DA content at P35.

Figure 4

Schematic of developmental photoperiod paradigm. For early adulthood midbrain RTPCR 5-HT gene experiments animals developed under either an (A) Equinox, (B) Long, or (C) Short photoperiod. For early adulthood DRN RNAScope experiments animals developed under either a (B) Long or (C) Short photoperiod. Animals developed under Equinox LD 12:12, Long LD 16:8, or Short LD 8:16 photoperiods from embryonic day 0 (E0) to postnatal day 50 (P50).

Figure 5

Long photoperiodic conditions resulted in significant decreases in expression levels of key 5-HT genes in the midbrain during early adulthood. (A) Tph2 expression, (B) SERT expression, and (C) Pet-1 expression. Mice developed under either Equinox (Eq), Long (L), or Short (S) photoperiods and 5-HT gene expression levels were measured at P50. The significant levels are as follows: (*p < 0.05, **p < 0.01). Note a trend level effect when comparing Short (S) and Long (L) photoperiods for Tph2 expression (A) (p = 0.0574). Reference gene for relative expression was Hprt.

Figure 6

RNAScope revealed significant differences in adult gene expression of relevant 5-HT genes in the DRN due to developmental photoperiod. Representative image for an animal that developed under a Long photoperiod and measured at P50: (A) Tph2 expression is stained in green and tagged with Alexa488, (B) SERT expression is stained in red and tagged with Atto550, (C) Pet-1 expression is stained in blue and tagged with Atto647, and (D) is the overlay of all three channels. Representative image for an animal that developed under a Short photoperiod and measured at P50: (E) Tph2 expression is stained in green and tagged with Alexa488, (F) SERT expression is stained in red and tagged with Atto550, (G) Pet-1 expression is stained in blue and tagged with Atto647, and (H) is the overlay of all three channels. Note that qualitatively, fluorescent levels are brighter under Short compared to Long photoperiod conditions. Scale bar was set at 100 μm.

Figure 7

Tph2 expression levels were significantly decreased in the DRN of mice developed under Long photoperiod conditions in early adulthood. (A) Integrated density of cell fluorescence, (B) integrated density of ROI fluorescence, (C) quantification of mean cell fluorescence, and (D) quantification of ROI mean fluorescence. Mice developed under either Long (L) or Short (S) photoperiods and RNAScope experiments occurred at P50. The significant levels are as follows: (*p < 0.05). Note a trend level effect when comparing Short and Long photoperiods for Integrated density of ROI fluorescence (B) (p = 0.0616).