Fig. 6. Impaired sympathetic innervation and altered AANAT induction in pineal glands from Egr3^-/- mice. The pineal glands from (A) wild-type and (A') Egr3^-/- mice appeared histologically indistinguishable. Immunohistochemistry for TH showed robust sympathetic innervation to the (B) wild-type pineal glands that was either highly diminished or absent in most pineal glands from Egr3^-/- mice (B'). Similar results were obtained with the lacZ sympathetic reporter that showed a robust network of sympathetic axons in (C) wild-type pineal glands that was barely detectable in (C') pineal glands from most Egr3^-/- mice. (D) AANAT is induced by increased sympathetic activity to the pineal upon entry into the dark phase of the light-dark cycle. In 10D:14L light cycle entrained wild-type mice, AANAT expression was induced as expected during the dark phase of the light-dark cycle and returned to low basal levels when sympathetic activity to the pineal gland decreased with the onset of the light phase of the cycle. However, in Egr3^-/- pineal glands AANAT was induced to 18% of the wild-type level, consistent with a physiological impairment of sympathetic innervation to the gland. The white and black bar along the horizontal axis depicts the time during the light-dark cycle when the lights are on and off, respectively. Zeitgeber time (ZT) represents the 24 hour elapsed time in the light-dark cycle relative to the time of dark-light transition (ZT0). (AANAT expression representative of three independent experiments and qPCR performed in triplicate; the peak wild-type value at ZT21 was considered 100% maximal AANAT induction in this paradigm; scale bars: 25 µm.)