Lateralization of circadian pacemaker output: Activation of left- and right-sided luteinizing hormone-releasing hormone neurons involves a neural rather than a humoral pathway

Abstract

Locomotor activity and luteinizing hormone (LH) secretion in golden hamsters share a common circadian pacemaker in the suprachiasmatic nucleus (SCN), but the rhythms do not seem to share a common output pathway from the SCN. Locomotion is believed to be driven by humoral factor(s), whereas LH secretion may depend on specific ipsilateral neural efferents from the SCN to LH releasing hormone (LHRH)-containing neurons in the preoptic area. In this paper we provide the first functional evidence for such efferents in neurologically intact hamsters by exploiting a phenomenon known as "splitting" in constant light, in which circa-12 hr (approximately 12 hr) locomotor activity bouts reflect an antiphase oscillation of the left and right sides of the bilaterally paired SCN. In ovariectomized, estrogen-treated (OVX + E2) female hamsters, splitting is also known to include circa-12 hr LH secretory surges. Here we show that behaviorally "split" OVX + E2 females exhibit a marked left-right asymmetry in immunoreactive c-Fos expression in both SCN and activated LHRH neurons, with the percentage of LHRH+/c-Fos+ double-labeled cells approximately fivefold higher on the side corresponding to the side of the SCN with higher c-Fos immunoreactivity. Our results suggest that splitting involves alternating left- and right-sided stimulation of LHRH neurons; under such circumstances, the functional activity of the neuroendocrine hypothalamus mirrors intrinsic side-to-side differences in SCN gene expression. The circadian regulation of reproductive activity depends on lateralized, point-to-point axonal projections rather than on diffusible factors.

Figure 1.

Behavioral splitting and immunoreactive c-Fos expression in the SCN. Left, Wheel-running activity of a representative female hamster maintained in LL, with the number of wheel revolutions over the course of each 24 hr period charted horizontally from left to right and succeeding days stacked vertically from top to bottom. At the white arrow, the animal's single daily bout of activity dissociated into two split components stably coupled ∼12 hr apart (∼30% of females exposed to LL exhibited splitting within 90 d). The animal was then ovariectomized (OVX), subcutaneously implanted with an estradiol benzoate capsule 14 d later (E2), and killed after 2 d (asterisk); 20% of the females, whether split or nonsplit, had disrupted wheel-running rhythms after OVX and were discarded from the study. Right, Coronal brain section through the SCN of a behaviorally split female hamster, killed 1 hr before the onset of a split activity bout and processed for c-Fos immunohistochemistry. 3V, Third ventricle; OC, optic chiasm. Scale bar, 500 μm.

Figure 2.

Asymmetric activation of LHRH-containing neurons during splitting. LHRH⁺ (blue) single-labeled cells (white arrows, left) and LHRH⁺ (blue)/c-Fos⁺ (brown) double-labeled cells (black arrows, right) from the left and right sides (red squares in insets) of the brain of the hamster whose SCN is shown in Figure 1. Similar results were observed using confocal microscopy (0.5-μm-thick optical sections) and immunofluorescence (data not shown). Scale bar, 50 μm.