Aging affects GABAergic function and calcium homeostasis in the mammalian central clock.

Introduction: Aging impairs the function of the central circadian clock in mammals, the suprachiasmatic nucleus (SCN), leading to a reduction in the output signal. The weaker timing signal from the SCN results in a decline in rhythm strength in many physiological functions, including sleep-wake patterns. Accumulating evidence suggests that the reduced amplitude of the SCN signal is caused by a decreased synchrony among the SCN neurons. The present study was aimed to investigate the hypothesis that the excitation/inhibition (E/I) balance plays a role in synchronization within the network. Methods: Using calcium (Ca2+) imaging, the polarity of Ca2+ transients in response to GABA stimulation in SCN slices of old mice (20-24 months) and young controls was studied. Results: We found that the amount of GABAergic excitation was increased, and that concordantly the E/I balance was higher in SCN slices of old mice when compared to young controls. Moreover, we showed an effect of aging on the baseline intracellular Ca2+ concentration, with higher Ca2+ levels in SCN neurons of old mice, indicating an alteration in Ca2+ homeostasis in the aged SCN. We conclude that the change in GABAergic function, and possibly the Ca2+ homeostasis, in SCN neurons may contribute to the altered synchrony within the aged SCN network.

Single cell model for re-entrainment to a shifted light cycle.

Our daily 24-h rhythm is synchronized to the external light-dark cycle resulting from the Earth's daily rotation. In the mammalian brain, the suprachiasmatic nucleus (SCN) serves as the master clock and receives light-mediated input via the retinohypothalamic tract. Abrupt changes in the timing of the light-dark cycle (e.g., due to jet lag) cause a phase shift in the circadian rhythms in the SCN. Here, we investigated the effects of a 6-h delay in the light-dark cycle on PERIOD2::LUCIFERASE expression at the single-cell level in mouse SCN organotypic explants. The ensemble pattern in phase shift response obtained from individual neurons in the anterior and central SCN revealed a bimodal distribution; specifically, neurons in the ventrolateral SCN responded with a rapid phase shift, while neurons in the dorsal SCN generally did not respond to the shift in the light-dark cycle. We also stimulated the hypothalamic tract in acute SCN slices to simulate light-mediated input to the SCN; interestingly, we found similarities between the distribution and fraction of rapid shifting neurons (in response to the delay) and neurons that were excited in response to electrical stimulation. These results suggest that a subpopulation of neurons in the ventral SCN that have an excitatory response to light input, shift their clock more readily than dorsal located neurons, and initiate the SCN's entrainment to the new light-dark cycle. Thus, we propose that light-excited neurons in the anterior and central SCN play an important role in the organism's ability to adjust to changes in the external light-dark cycle.

Inhibitory responses to retinohypothalamic tract stimulation in the circadian clock of the diurnal rodent Rhabdomys pumilio.

In both diurnal and nocturnal mammals, the timing of activity is regulated by the central circadian clock of the suprachiasmatic nucleus (SCN). The SCN is synchronized to the external light cycle via the retinohypothalamic tract (RHT). To investigate potential differences in light processing between nocturnal mice and the diurnal rodent Rhabdomys pumilio, we mimicked retinal input by stimulation of the RHT ex vivo. Using Ca2+ imaging, we observed excitations as well as inhibitions of SCN neurons in response to electrical RHT stimulation. In mice, the vast majority of responses were excitatory (85%), whereas in Rhabdomys, the proportion of excitatory and inhibitory responses was similar (51% excitatory, 49% inhibitory). Glutamate blockers AP5 and CNQX blocked the excitatory responses to RHT stimulation but did not abolish the inhibitory responses in mice or Rhabdomys, indicating that the inhibitions were monosynaptically transmitted via the RHT. Simultaneous application of glutamate blockers with the GABAA antagonist gabazine blocked all inhibitory responses in mice, but not in Rhabdomys. Collectively, our results indicate that in Rhabdomys, considerably more inhibitory responses to light are present and that these responses are driven directly by the RHT. We propose that this increased proportion of inhibitory input could reflect a difference in the entrainment mechanism employed by diurnal rodents.