Michael Menaker

The Intergeniculate Leaflet Partially Mediates Effects of Light on Circadian Rhythms

Photic signals affect circadian activity rhythms by both phasic and tonic mechanisms that modulate pacemaker phase and period. In mammals, the effects of light on circadian activity are mediated by the retina, which communicates with the suprahiasmatic nucleus (SCN) by two different anatomical routes: the retino-hypothalamic tract (RHT), originating in the retina, and the geniculo-hypothalamic tract (GHT), arising from a retino-recipient nucleus, the intergeniculate leaflet (IGL). We assessed the roles of these two afferent systems in mediating phasic and tonic effects of light on circadian activity in IGL-lesioned animals. Destruction of the IGL significantly affected phase shifts produced by brief light pulses (phasic effect) and modified the change in period (τ) of the free-running activity rhythm produced by changing the level of constant light (LL) (tonic effect). Phase advances produced by brief light pulses were decreased in amplitude while phase delays were increased in IGL-lesioned animals as compared to controls. The free-running period in constant dark (τDD) of IGL-lesioned animals was greater than τDD of controls, and the lengthening of τ normally produced by LL was not observed or was greatly reduced in IGL-lesioned animals. Entrainment to light-dark cycles was unaffected by the lesions, as were other aspects of the circadian activity rhythm that normally change in response to LL (e.g., activity-rest ratio, total activity, splitting). Our data support the interpretation that the IGL plays a significant role in relaying information regarding illu mination intensity to the SCN.

Dose-response relationship between light irradiance and the suppression of plasma melatonin in human volunteers

This study tested the capacity of different irradiances of monochromatic light to reduce plasma melatonin in normal humans. Six healthy male volunteers, 24-34 years old, were exposed to 0.01, 0.3, 1.6, 5, or 13 microW/cm2 of 509 nm monochromatic light for 1 h during the night on separate occasions. Light irradiance depressed plasma melatonin in a dose-response pattern. The data indicate that the mean threshold irradiance for suppressing melatonin is between 1.6 and 5 microW/cm2. Individual variations in threshold responses to monochromatic light were observed among the volunteers.

Bicuculline blocks circadian phase delays but not advances

Phase delays of hamster locomotor rhythms, which normally can be produced by light pulses given in the early subjective night, were blocked by bicuculline, a selective antagonist of gamma-aminobutyric acid (GABA) activity. Phase advances induced by light pulses given in the late subjective night were unaffected by bicuculline. This suggests that GABA may be involved in the mediation of some, but not all, light input to the mammalian circadian system. Furthermore, it raises the possibility that light input may be mediated at different circadian time points by functionally separate pathways.

Effects of diazepam on circadian phase advances and delays

Phase advances of hamster locomotor rhythms, which normally can be induced by light pulses in the late subjective night, were blocked in a dose-dependent manner by the benzodiazepine, diazepam. Light-induced phase delays were unaffected at doses that significantly blocked phase advances. Diazepam caused small phase delays of the free-running rhythm when given without a light pulse at either phase advance or phase delay time points. These results are discussed with regard to the possibility that different neurochemical mechanisms are required to process light-induced phase advances and delays and that GABA neurotransmission may be involved in the modulation of light input to the clock.

Effects of excitatory amino acid receptor antagonists and agonists on suprachiasmatic nucleus responses to retinohypothalamic tract volleys.

A slice preparation of the mouse hypothalamus that includes the suprachiasmatic nuclei (SCN), the optic chiasm and the optic nerves was used for pharmacologic investigations of the nature of the receptors mediating the excitation of SCN neurons by input from the retinohypothalamic tract (RHT). Bath application of cis-2,3-piperidinedicarboxylic acid, a non-selective antagonist of excitatory amino acid receptors, reversibly blocked the postsynaptic component of the field potentials evoked in the dorsolateral SCN by stimulation of the optic nerve. The selective antagonist of N-methyl-D-aspartate receptors, 2-amino-5-phosphonovaleric acid, had no effect on SCN responses. Glutamic acid diethyl ester and 2-amino-4-phosphonobutyric acid also were without effect, but gamma-D-glutamylglycine caused a small decrease in the amplitude of the postsynaptic wave. Addition of the agonists, kainate and N-methyl-D,L-aspartate, to the superfusate also blocked the postsynaptic response. Kainate was the most potent agonist. L-Glutamate was without effect at up to 100 microM. These results indicate that postsynaptic responses in the SCN to retinohypothalamic tract volleys are mediated by a non-NMDA class of excitatory amino acid receptors.

Sympathetic regulation of chicken pineal rhythms

Adult hens were chronically cannulated and held in light-dark (LD) 12:12 h lighting regimes or in constant darknesS (DD). Periodic blood sampling for 5-9 days revealed circadian rhythms in plasma melatonin titres. Superior cervical ganglionectomy (SCG-X) performed 1 week after hatching had little or no effect on these rhythms in LD, but unlike normals. SCG-X birds did not sustain persistent rhythms in DD. In SCG-X birds, norepinephrine (NE) infusion for 12 h of each 24 h in DD significantly reduced plasma melatonin titres during the infusion and re-established a rhythm. After each experiment, hens were killed, their pineals were removed and assayed by HPLC-EC for NE, dopamine (DA), serotonin (5-HT) and 5-hydroxy-3-indole-acetic acid (5-HIAA). SCG-X resulted in a 90% depletion of pineal NE: DA content was reduced to undetectable levels. Pineal 5-HT and 5-HIAA were also reduced by SCG-X. The chicken pineal contains circadian oscillators which persist in vitro8.19.29. The results reported here suggest that noradrenergic fibres from the SCG regulate the pineals inherent rhythmicity. NE normally released from sympathetic terminals during the birds day may synchronize oscillators within the pineal by inhibiting melatonin synthesis.

Light-induced decrease of serotonin N-acetyltransferase activity and melatonin in the chicken pineal gland and retina

In the pineal gland and retina of chickens, light exposure at night when serotonin N-acetyltransferase (NAT) activity levels are high causes a 4--5-fold decrease in NAT activity. The t1/2 of NAT inactivation is 10 min and the kinetics of inactivation are similar in the pineal gland and retina. A brief pulse of light can initiate the process of NAT inactivation which continues in the dark for 30 min before a partial recovery of NAT activity occurs. In blinded chicks, there is less inactivation of pineal NAT by light than in intact chicks, indicating that the eyes are involved in the pineals response to light. In chicks that have had their superior cervical ganglia removed inactivation of pineal NAT by light is intermediate between that of intact and blind chicks, indicating that ganglionectomy does not completely mimic the effects of blinding. The pineal gland itself is light-sensitive in culture. Light causes a 4--5-fold decrease in NAT activity in static organ culture, and inhibits melatonin release in flow-through organ culture. Drugs that increase cyclic nucleotide levels in cells (cholera toxin, RO 20-1724, monobutyryl cyclic AMP, monobutyryl cyclic GMP) block the NAT decrease by light, whereas high potassium or EGTA do not block this light-induced NAT inactivation.

Multiple redundant circadian oscillators within the isolated avian pineal gland

SummaryThe avian pineal gland contains a circadian pacemaker that oscillates in vitro. Using a flow-through culture system it is possible to measure melatonin production from very small subsections of an individual gland. We have used this technique to attempt to localize the oscillators in the pineal. Progressive tissue reduction did not affect the rhythmicity of cultured pineals. Multiple pieces (up to eight) from a single pineal all were capable of circadian oscillation — establishing directly that a pineal gland contains at least eight oscillators. All pineal pieces were responsive to light, and single light pulses shifted the phase of the melatonin rhythm. Because pieces equivalent to less than one per cent of the whole gland were rhythmic and because the capacity for oscillation was distributed throughout the gland, an individual pineal appears to be composed of a population of circadian oscillators.

Kynurenic acid blocks suprachiasmatic nucleus responses to optic nerve stimulation.

An in vitro slice preparation of the mouse hypothalamus was used to determine the effects of pharmacological agents on the field potentials that are evoked in the suprachiasmatic nucleus (SCN) by stimulation of the optic nerve. Postsynaptic components of these responses were identified by lowering the concentration of calcium in the superfusate. Bath application of kynurenate, an antagonist of excitatory amino acid neurotransmission, reversibly blocked postsynaptic responses in the SCN. The evoked responses in the SCN were not affected by the acetylcholinergic agents (+)-tubocurarine, scopolamine, physostigmine, or carbachol. These results suggest that excitatory amino acid receptors mediate responses of SCN neurons to retinal input, but do not support a role for acetylcholine.

Circadian locomotor rhythms in the desert iguana I. The role of the eyes and the pineal

SummaryThe pineal and the eyes are known to be important components in the circadian system of some species of lizards; their effects may be mediated by the hormone melatonin. We examined the role played by these structures in the desert iguana (Dipsosaurus dorsalis). Surgical removal of the pineal had no effect on circadian locomotor rhythms, even though this procedure abolished the circadian rhythm of melatonin in the blood. Furthermore, when the isolated pineal of Dipsosaurus was studied in organ culture, it showed no circadian rhythm of melatonin secretion, as do pineals of some other lizard species, although it did produce large quantities of this hormone. Bilateral ocular enucleation had only small effects on the freerunning period of locomotor rhythms, without affecting melatonin levels in the blood. Behavioral circadian rhythms persisted in desert iguanas subjected to both enucleation and pinealectomy. These data suggest that neither the pineal nor the eyes are central components of the circadian pacemaking system in Dipsosaurus, nor is melatonin critically involved in maintaining its organization.