Daily peak plasma glucose concentrations are attained shortly before awakening. Previous experiments indicated an important role for the biological clock, located in the suprachiasmatic nuclei (SCN), in the genesis of this anticipatory rise in plasma glucose concentrations by controlling hepatic glucose production. Here, we show that stimulation of NMDA receptors, or blockade of GABA receptors in the paraventricular nucleus of the hypothalamus (PVN) of conscious rats, caused a pronounced increase in plasma glucose concentrations. The local administration of TTX in brain areas afferent to the PVN revealed that an important part of the inhibitory inputs to the PVN was derived from the SCN. Using a transneuronal viral-tracing technique, we showed that the SCN is connected to the liver via both branches of the autonomic nervous system (ANS). The combination of a blockade of GABA receptors in the PVN with selective removal of either the sympathetic or parasympathetic branch of the hepatic ANS innervation showed that hyperglycemia produced by PVN stimulation was primarily attributable to an activation of the sympathetic input to the liver. We propose that the daily rise in plasma glucose concentrations is caused by an SCN-mediated withdrawal of GABAergic inputs to sympathetic preautonomic neurons in the PVN, resulting in an increased hepatic glucose production. The remarkable resemblance of the presently proposed control mechanism to that described previously for the control of daily melatonin rhythm suggests that the GABAergic control of sympathetic preautonomic neurons in the PVN is an important pathway for the SCN to control peripheral physiology.

In order to drive tissue-specific rhythmic outputs, the master clock, located in the suprachiasmatic nucleus (SCN), is thought to reset peripheral oscillators via either chemical and hormonal cues or neural connections. Recently, the daily rhythm of plasma glucose (characterized by a peak before the onset of the activity period) has been shown to be directly driven by the SCN, independently of the SCN control of rhythmic feeding behaviour. Indeed, the daily variation in glucose was not impaired unless the scheduled feeding regimen (six-meal schedule) was associated with an SCN lesion. Here we show that the rhythmicity of both clock-gene mRNA expression in the liver and plasma glucose is not abolished under such a regular feeding schedule. Because the onset of the activity period and hyperglycemia are correlated with an increased sympathetic tonus, we investigated whether this autonomic branch is involved in the SCN control of plasma glucose rhythm and liver rhythmicity. Interestingly, hepatic sympathectomy combined with a six-meal feeding schedule resulted in a disruption of the plasma glucose rhythmicity without affecting the daily variation in clock-gene mRNA expression in the liver. Taking all these data together, we conclude that (i) the SCN needs the sympathetic pathway to the liver to generate the 24-h rhythm in plasma glucose concentrations, (ii) rhythmic clock-gene expression in the liver is not dependent on the sympathetic liver innervation and (iii) clock-gene rhythmicity in liver cells is not sufficient for sustaining a circadian rhythm in plasma glucose concentrations.

The suprachiasmatic nucleus (SCN) controls the circadian rhythm of melatonin synthesis in the mammalian pineal gland by a multisynaptic pathway including, successively, preautonomic neurons of the paraventricular nucleus (PVN), sympathetic preganglionic neurons in the spinal cord and noradrenergic neurons of the superior cervical ganglion (SCG). In order to clarify the role of each of these structures in the generation of the melatonin synthesis rhythm, we first investigated the day- and night-time capacity of the rat pineal gland to produce melatonin after bilateral SCN lesions, PVN lesions or SCG removal, by measurements of arylalkylamine N-acetyltransferase (AA-NAT) gene expression and pineal melatonin content. In addition, we followed the endogenous 48 h-pattern of melatonin secretion in SCN-lesioned vs. intact rats, by microdialysis in the pineal gland. Corticosterone content was measured in the same dialysates to assess the SCN lesions effectiveness. All treatments completely eliminated the day/night difference in melatonin synthesis. In PVN-lesioned and ganglionectomised rats, AA-NAT levels and pineal melatonin content were low (i.e. 12% of night-time control levels) for both day- and night-time periods. In SCN-lesioned rats, AA-NAT levels were intermediate (i.e. 30% of night-time control levels) and the 48-h secretion of melatonin presented constant levels not exceeding 20% of night-time control levels. The present results show that ablation of the SCN not only removes an inhibitory input but also a stimulatory input to the melatonin rhythm generating system. Combination of inhibitory and stimulatory SCN outputs could be of a great interest for the mechanism of adaptation to day-length (i.e. adaptation to seasons).

Plasma glucose concentrations display a daily rhythm generated by the hypothalamic biological clock, located in the suprachiasmatic nucleus (SCN). How the SCN orchestrates this rhythm is unknown. Because glucagon stimulates hepatic glucose production, we hypothesized that if glucagon has a daily rhythm, then it may be responsible for the glucose rhythm. From hourly blood samples, we determined daily glucagon concentrations for intact and SCN-lesioned rats. Intact ad libitum-fed rats showed a clear daily glucagon rhythm, and fasting resulted in an even more pronounced rhythm. It is interesting that a decrease in glucagon concentrations, instead of the expected increase, occurred already shortly after food removal. Toward the start of the active period, a peak in glucagon levels occurred, with concentrations similar to those measured in ad libitumfed rats. SCN lesions abolished rhythmicity in plasma glucagon profiles. Scheduled-fed rats showed meal-induced glucagon peaks but also a daily rhythm in basal premeal glucagon concentrations. Plasma glucose concentrations of ad libitum-and scheduled-fed rats, however, were similar. In conclusion, feeding and the biological clock control 24-h plasma glucagon concentrations. In fed rats, glucagon is not responsible for the daily glucose rhythm. During fasting, however, glucagon may contribute to energy mobilization when the activity period starts. Diabetes 52: 1709 -1715, 2003 M ammals adapt their activity pattern to the daily changes in light intensity. This activity rhythm dictates the need for energy at a specific time of day. This includes glucose in particular, because under normal conditions, it is the only fuel that can be metabolized by the brain (1). Circadian rhythms have been shown in mammals (2,3), including humans (4,5). Rat plasma glucose concentrations display a daily rhythm, with peak values at the beginning of the activity period. Studies involving scheduled feeding and thermic lesions of the biological clock, located in the suprachiasmatic nucleus (SCN), have shown that this rhythm is feeding independent and generated by the SCN (6,7), yet it is not clear by which mechanism the SCN creates this glucose rhythm. Studies investigating the role of insulin have shown that insulin concentrations have a daily rhythm but not independent of food intake (6) and therefore cannot be responsible for the glucose rhythm. Corticosterone stimulates gluconeogenesis (8) and also displays a clear daily rhythm (9 -12), with peak values occurring just before the onset of the activity period. Despite the hyperglycemic effect of corticosterone and the coincidence of its peak release with the peak in plasma glucose concentrations, blocking corticosterone synthesis does not affect the morning rise of glucose concentrations seen in humans (13) and thus cannot be the main cause of the rhythm in glucose concentrations. Growth hormone, too, is able to stimulate glucose release (8) but in the rat displays an ultradian rhythm rather than a circadian rhythm, which makes it unlikely to...

The hypothalamus uses hormones and the autonomic nervous system to balance energy fluxes in the body. Here we show that the autonomic nervous system has a distinct organization in different body compartments. The same neurons control intraabdominal organs (intraabdominal fat, liver, and pancreas), whereas sc adipose tissue located outside the abdominal compartment receives input from another set of autonomic neurons. This differentiation persists up to preautonomic neurons in the hypothalamus, including the biological clock, that have a distinct organization depending on the body compartment they command. Moreover, we demonstrate a neuronal feedback from adipose tissue that reaches the brainstem. We propose that this compartment-specific organization offers a neuroanatomical perspective for the regional malfunction of organs in type 2 diabetes, where increased insulin secretion by the pancreas and disturbed glucose metabolism in the liver coincide with an augmented metabolic activity of visceral compared with sc adipose tissue.

The suprachiasmatic nucleus (SCN) is the "master clock" of the mammalian brain. It coordinates the peripheral clocks in the body, including the pineal clock that receives SCN input via a multisynaptic noradrenergic pathway. Rhythmic pineal melatonin production is disrupted in Alzheimer's disease (AD). Here we show that the clock genes hBmal1, hCry1, and hPer1 were rhythmically expressed in the pineal of controls (Braak 0). Moreover, hPer1 and hbeta1-adrenergic receptor (hbeta1-ADR) mRNA were positively correlated and showed a similar daily pattern. In contrast, in both preclinical (Braak I-II) and clinical AD patients (Braak V-VI), the rhythmic expression of clock genes was lost as well as the correlation between hPer1 and hbeta1-ADR mRNA. Intriguingly, hCry1 mRNA was increased in clinical AD. These changes are probably due to a disruption of the SCN control, as they were mirrored in the rat pineal deprived of SCN control. Indeed, a functional disruption of the SCN was observed from the earliest AD stages onward, as shown by decreased vasopressin mRNA, a clock-controlled major output of the SCN. Thus, a functional disconnection between the SCN and the pineal from the earliest AD stage onward could account for the pineal clock gene changes and underlie the circadian rhythm disturbances in AD.

BackgroundThe biological clock, located in the hypothalamic suprachiasmatic nucleus (SCN), controls the daily rhythms in physiology and behavior. Early studies demonstrated that light exposure not only affects the phase of the SCN but also the functional activity of peripheral organs. More recently it was shown that the same light stimulus induces immediate changes in clock gene expression in the pineal and adrenal, suggesting a role of peripheral clocks in the organ-specific output. In the present study, we further investigated the immediate effect of nocturnal light exposure on clock genes and metabolism-related genes in different organs of the rat. In addition, we investigated the role of the autonomic nervous system as a possible output pathway of the SCN to modify the activity of the liver after light exposure.Methodology and Principal FindingsFirst, we demonstrated that light, applied at different circadian times, affects clock gene expression in a different manner, depending on the time of day and the organ. However, the changes in clock gene expression did not correlate in a consistent manner with those of the output genes (i.e., genes involved in the functional output of an organ). Then, by selectively removing the autonomic innervation to the liver, we demonstrated that light affects liver gene expression not only via the hormonal pathway but also via the autonomic input.ConclusionNocturnal light immediately affects peripheral clock gene expression but without a clear correlation with organ-specific output genes, raising the question whether the peripheral clock plays a “decisive” role in the immediate (functional) response of an organ to nocturnal light exposure. Interestingly, the autonomic innervation of the liver is essential to transmit the light information from the SCN, indicating that the autonomic nervous system is an important gateway for the SCN to cause an immediate resetting of peripheral physiology after phase-shift inducing light exposures.

OBJECTIVE-We recently showed that intracerebroventricular infusion of neuropeptide Y (NPY) hampers inhibition of endogenous glucose production (EGP) by insulin in mice. The downstream mechanisms responsible for these effects of NPY remain to be elucidated. Therefore, the aim of this study was to establish whether intracerebroventricular NPY administration modulates the suppressive action of insulin on EGP via hepatic sympathetic or parasympathetic innervation. RESEARCH DESIGN AND METHODS-The effects of a continuous intracerebroventricular infusion of NPY on glucose turnover were determined in rats during a hyperinsulinemiceuglycemic clamp. Either rats were sham operated, or the liver was sympathetically (hepatic sympathectomy) or parasympathetically (hepatic parasympathectomy) denervated.RESULTS-Sympathectomy or parasympathectomy did not affect the capacity of insulin to suppress EGP in intracerebroventricular vehicle-infused animals (50 Ϯ 8 vs. 49 Ϯ 6 vs. 55 Ϯ 6%, in hepatic sympathectomy vs. hepatic parasympathectomy vs. sham, respectively). Intracerebroventricular infusion of NPY significantly hampered the suppression of EGP by insulin in sham-denervated animals (29 Ϯ 9 vs. 55 Ϯ 6% for NPY/sham vs. vehicle/sham, respectively, P ϭ 0.038). Selective sympathetic denervation of the liver completely blocked the effect of intracerebroventricular NPY administration on insulin action to suppress EGP (NPY/hepatic sympathectomy, 57 Ϯ 7%), whereas selective parasympathetic denervation had no effect (NPY/hepatic parasympathectomy, 29 Ϯ 7%).CONCLUSIONS-Intracerebroventricular administration of NPY acutely induces insulin resistance of EGP via activation of sympathetic output to the liver.