The related neuropeptides vasoactive intestinal peptide (VIP) and peptide histidine isoleucine (PHI) are expressed at high levels in the neurons of the suprachiasmatic nucleus (SCN), but their function in the regulation of circadian rhythms is unknown. To study the role of these peptides on the circadian system in vivo, a new mouse model was developed in which both VIP and PHI genes were disrupted by homologous recombination. In a light-dark cycle, these mice exhibited diurnal rhythms in activity which were largely indistinguishable from wild-type controls. In constant darkness, the VIP/PHI-deficient mice exhibited pronounced abnormalities in their circadian system. The activity patterns started approximately 8 h earlier than predicted by the previous light cycle. In addition, lack of VIP/PHI led to a shortened free-running period and a loss of the coherence and precision of the circadian locomotor activity rhythm. In about one-quarter of VIP/PHI mice examined, the wheel-running rhythm became arrhythmic after several weeks in constant darkness. Another striking example of these deficits is seen in the split-activity patterns expressed by the mutant mice when they were exposed to a skeleton photoperiod. In addition, the VIP/PHI-deficient mice exhibited deficits in the response of their circadian system to light. Electrophysiological analysis indicates that VIP enhances inhibitory synaptic transmission within the SCN of wild-type and VIP/PHI-deficient mice. Together, the observations suggest that VIP/PHI peptides are critically involved in both the generation of circadian oscillations as well as the normal synchronization of these rhythms to light.

The present experiments were designed to examine dopamine (DA) modulation of whole cell currents mediated by activation of N-methyl-D-aspartate (NMDA) receptors in visualized neostriatal neurons in slices. First, we assessed the ability of DA, D1 and D2 receptor agonists to modulate membrane currents induced by activation of NMDA receptors. The results of these experiments demonstrated that DA potentiated NMDA-induced currents in medium-sized neostriatal neurons. Potentiation of NMDA currents occurred at three different holding potentials, although it was more pronounced at -30 mV. It was mediated by D1 receptors, because it was mimicked by D1 agonists and blocked by exposure to a D1 antagonist. Activation of D2 receptors produced inconsistent effects on NMDA-induced membrane currents. Either decreases, increases, or no effects on NMDA currents occurred. Second, we examined the contributions of intrinsic, voltage-dependent conductances to DA potentiation of NMDA currents. Blockade of K+ conductances did not prevent DA enhancement of NMDA currents. However, voltage-activated Ca2+ conductances provided a major contribution to DA modulation. The dihydropyridine L-type Ca2+ channel blockers, nifedipine, and methoxyverapamil (D-600), markedly reduced but did not totally eliminate the ability of DA to modulate NMDA currents. The D1 receptor agonist SKF 38393 also enhanced Ba2+ currents in neostriatal neurons. Together, these findings provide evidence for a complex interplay between DA, NMDA receptor activation and dihydropyridine-sensitive Ca2+ conductances in controlling responsiveness of neostriatal medium-sized neurons.

In mammals, the part of the nervous system responsible for most circadian behavior can be localized to a pair of structures in the hypothalamus known as the suprachiasmatic nucleus (SCN). Previous studies suggest that the basic mechanism responsible for the generation of these rhythms is intrinsic to individual cells. There is also evidence that the cells within the SCN are coupled to one another and that this coupling is important for the normal functioning of the circadian system. One mechanism that mediates coordinated electrical activity is direct electrical connections between cells formed by gap junctions. In the present study, we used a brain slice preparation to show that developing SCN cells are dye coupled. Dye coupling was observed in both the ventrolateral and dorsomedial subdivisions of the SCN and was blocked by application of a gap junction inhibitor, halothane. Dye coupling in the SCN ap-pears to be regulated by activity-dependent mechanisms as both tetrodotoxin and the GABA A agonist muscimol inhibited the extent of coupling. Furthermore, acute hyperpolarization of the membrane potential of the original biocytin-filled neuron decreased the extent of coupling. SCN cells were extensively dye coupled during the day when the cells exhibit synchronous neural activity but were minimally dye coupled during the night when the cells are electrically silent. Immunocytochemical analysis provides evidence that a gap-junction-forming protein, connexin32, is expressed in the SCN of postnatal animals. Together the results are consistent with a model in which gap junctions provide a means to couple SCN neurons on a circadian basis.

The purpose of the present study was to examine whether cAMP-dependent mechanisms regulated excitatory synaptic transmission in the neostriatum. A brain slice preparation was utilized for intracellular recordings of the excitatory postsynaptic potentials (EPSPs) evoked by electrical stimulation. Bath application of forskolin, an activator of adenylate cyclase, enhanced the EPSP amplitude and duration. This potentiation was dose dependent and did not occur with the inactive analog 1,9-dideoxyforskolin. Forskolin potentiation was unaltered by treatment with the GABAA receptor antagonist bicuculline. Furthermore, two inhibitors of cAMP-dependent protein kinase (PKA), Rp-cAMPS and IP20-amide, attenuated forskolin's enhancement of the EPSP. In addition, the PKA activator Sp-cAMPS enhanced excitatory synaptic transmission. Interestingly, treatment with PKA inhibitors alone depressed while the phosphatase inhibitor okadaic acid enhanced the synaptic response. These results suggest a role for tonic kinase and phosphatase activity in regulating excitatory synaptic transmission in the neostriatum. Finally, forskolin was found to enhance the responses of neostriatal neurons to glutamate receptor agonists. This potentiation, which occurred in the presence of tetrodotoxin, provides at leas part of the explanation for the cAMP/PKA-dependent regulation of the EPSP. Overall, these results suggest a role for the adenylate cyclase cascade in the regulation of excitatory synaptic transmission in the neostriatum.

Circadian timing systems, like most physiological processes, cannot escape the effects of aging. With age, humans experience decreased duration and quality of sleep. Aged mice exhibit decreased amplitude and increased fragmentation of the activity rhythm, and lengthened circadian free-running period in both light-dark (LD) and constant dark (DD) conditions. Several studies have shown that aging impacts neural activity rhythms in the central circadian clock in the suprachiasmatic nucleus (SCN). However, evidence for age-related disruption of circadian oscillations of clock genes in the SCN has been equivocal. We hypothesized that daily exposure to LD cycles masks the full impact of aging on molecular rhythms in the SCN. We performed ex vivo bioluminescent imaging of cultured SCN slices of young and aged PER2::luciferase knock-in (PER2::LUC) mice housed under LD or prolonged DD conditions. Under LD conditions, the amplitude of PER2::LUC rhythms differed only slightly between SCN explants from young and aged animals; under DD conditions, the PER2::LUC rhythms of aged animals showed markedly lower amplitudes and longer circadian periods than those of young animals. Recordings of PER2::LUC rhythms in individual SCN cells using an electron multiplying charge-coupled device camera revealed that aged SCN cells showed longer circadian periods and that the rhythms of individual cells rapidly became desynchronized. These data suggest that aging degrades the SCN circadian ensemble, but that recurrent LD cycles mask these effects. We propose that these changes reflect a decline in pacemaker robustness that could increase vulnerability to environmental challenges, and partly explain age-related sleep and circadian disturbances.

A variety of evidence suggests that the effects of light on the mammalian circadian system are mediated by direct retinal ganglion cell projection to the suprachiasmatic nucleus (SCN). This synaptic connection is glutamatergic and the release of glutamate is detected by both N-methyl-D-asparate (NMDA) and amino-methyl proprionic acid/kainate (AMPA/KA) iontotropic glutamate receptors (GluRs). It is well established that NMDA GluRs play a critical role in mediating the effects of light on the circadian system; however, the role of AMPA/KA GluRs has received less attention. In the present study, we sought to better understand the contribution of AMPA/KA-mediated currents in the circadian system based in the SCN. First, whole cell patch-clamp electrophysiological techniques were utilized to measure spontaneous excitatory postsynaptic currents (sEPSCs) from SCN neurons. These currents were widespread in the SCN and not just restricted to the retino-recipient region. The sEPSC frequency and amplitude did not vary with the daily cycle. Similarly, currents evoked by the exogenous application of AMPA onto SCN neurons were widespread within the SCN and did not exhibit a diurnal rhythm in their magnitude. Fluorometric techniques were utilized to estimate AMPA-induced calcium (Ca(2+)) concentration changes in SCN neurons. The resulting data indicate that AMPA-evoked Ca(2+) transients were widespread in the SCN and that there was a daily rhythm in the magnitude of AMPA-induced Ca(2+) transients that peaked during the night. By itself, blocking AMPA/KA GluRs with a receptor blocker decreased the spontaneous firing of some SCN neurons as well as reduced resting Ca(2+) levels, suggesting tonic glutamatergic excitation. Finally, immunohistochemical techniques were used to describe expression of the AMPA-preferring GluR subunits GluR1 and GluR2/3s within the SCN. Overall, our data suggest that glutamatergic synaptic transmission mediated by AMPA/KA GluRs play an important role throughout the SCN synaptic circuitry.

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Objective To review and discuss the putative role of light, sleep, and the biological clock in cluster headache. Discussion Cluster headache attacks are believed to be modulated in the hypothalamus; moreover, the severe pain and typical autonomic cranial features associated with cluster headache are caused by abnormal activity of the trigeminal-autonomic reflex. The temporal pattern of cluster headache attacks suggests involvement of the biological clock, and the seasonal pattern is influenced by the number of daylight hours. Although sleep is often reported as a trigger for cluster headache attacks, to date no clear correlation has been established between these attacks and sleep stage. Conclusions We hypothesize that light, sleep, and the biological clock can change the brain’s state, thereby lowering the threshold for activating the trigeminal-autonomic reflex, resulting in a cluster headache attack. Understanding the mechanisms that contribute to the daily and seasonal fluctuations in cluster headache attacks may provide new therapeutic targets.