Increased ghrelin signaling prolongs survival in mouse models of human aging through activation of sirtuin1.

Caloric restriction (CR) is known to retard aging and delay functional decline as well as the onset of diseases in most organisms. Ghrelin is secreted from the stomach in response to CR and regulates energy metabolism. We hypothesized that in CR ghrelin has a role in protecting aging-related diseases. We examined the physiological mechanisms underlying the ghrelin system during the aging process in three mouse strains with different genetic and biochemical backgrounds as animal models of accelerated or normal human aging. The elevated plasma ghrelin concentration was observed in both klotho-deficient and senescence-accelerated mouse prone/8 (SAMP8) mice. Ghrelin treatment failed to stimulate appetite and prolong survival in klotho-deficient mice, suggesting the existence of ghrelin resistance in the process of aging. However, ghrelin antagonist hastened death and ghrelin signaling potentiators rikkunshito and atractylodin ameliorated several age-related diseases with decreased microglial activation in the brain and prolonged survival in klotho-deficient, SAMP8 and aged ICR mice. In vitro experiments, the elevated sirtuin1 (SIRT1) activity and protein expression through the cAMP-CREB pathway was observed after ghrelin and ghrelin potentiator treatment in ghrelin receptor 1a-expressing cells and human umbilical vein endothelial cells. Furthermore, rikkunshito increased hypothalamic SIRT1 activity and SIRT1 protein expression of the heart in the all three mouse models of aging. Pericarditis, myocardial calcification and atrophy of myocardial and muscle fiber were improved by treatment with rikkunshito. Ghrelin signaling may represent one of the mechanisms activated by CR, and potentiating ghrelin signaling may be useful to extend health and lifespan.

Potentiation of ghrelin signaling attenuates cancer anorexia-cachexia and prolongs survival.

Cancer anorexia-cachexia syndrome is characterized by decreased food intake, weight loss, muscle tissue wasting and psychological distress, and this syndrome is a major source of increased morbidity and mortality in cancer patients. This study aimed to clarify the gut-brain peptides involved in the pathogenesis of the syndrome and determine effective treatment for cancer anorexia-cachexia. We show that both ghrelin insufficiency and resistance were observed in tumor-bearing rats. Corticotropin-releasing factor (CRF) decreased the plasma level of acyl ghrelin, and its receptor antagonist, α-helical CRF, increased food intake of these rats. The serotonin 2c receptor (5-HT2cR) antagonist SB242084 decreased hypothalamic CRF level and improved anorexia, gastrointestinal (GI) dysmotility and body weight loss. The ghrelin receptor antagonist (D-Lys3)-GHRP-6 worsened anorexia and hastened death in tumor-bearing rats. Ghrelin attenuated anorexia-cachexia in the short term, but failed to prolong survival, as did SB242084 administration. In addition, the herbal medicine rikkunshito improved anorexia, GI dysmotility, muscle wasting, and anxiety-related behavior and prolonged survival in animals and patients with cancer. The appetite-stimulating effect of rikkunshito was blocked by (D-Lys3)-GHRP-6. Active components of rikkunshito, hesperidin and atractylodin, potentiated ghrelin secretion and receptor signaling, respectively, and atractylodin prolonged survival in tumor-bearing rats. Our study demonstrates that the integrated mechanism underlying cancer anorexia-cachexia involves lowered ghrelin signaling due to excessive hypothalamic interactions of 5-HT with CRF through the 5-HT2cR. Potentiation of ghrelin receptor signaling may be an attractive treatment for anorexia, muscle wasting and prolong survival in patients with cancer anorexia-cachexia.

Hepato-vagal pathway associated with nicotine's anorectic effect in the rat.

Nicotine reduces appetite and body weight. Because the hepato-portal area senses different types of nutrients that transmit signals via vagal afferent nerves to the hypothalamus to modify food intake and feeding pattern, we investigated the effect of nicotine on a hepato-vagal-hypothalamic pathway. Low doses of nicotine (< 10 ng) injected into portal vein (i.p.v.) decreased, while high doses of nicotine increased (> or = 10 ng) electrophysiological activity of hepatic vagal afferents. Stimulatory effect of high dose of nicotine on vagal hepatic afferents was blocked by a prior i.p.v. injection of curare (30 microg) or hexamethonium (1 mg). Furthermore, activities of gastric vagal and adrenal sympathetic efferents were suppressed by low-dose, but stimulated by high-dose i.p.v. nicotine. These reflex effects did not occur in hepatic vagotomized rats. Results of experiments demonstrate that in addition to nicotine's anorectic effect being mediated via a direct central action, nicotine also acts peripherally via hepatic vagal afferents from sensors of nicotine in the hepato-portal region.

Parasympathetic and sympathetic control of the pancreas: a role for the suprachiasmatic nucleus and other hypothalamic centers that are involved in the regulation of food intake.

To reveal brain regions and transmitter systems involved in control of pancreatic hormone secretion, specific vagal and sympathetic denervation were combined with injection of a retrograde transsynaptic tracer, pseudorabies virus (PRV), into the pancreas. After sympathetic or vagal transsection first-order neurons were revealed in the dorsal motor nucleus of the vagus (DMV) or in preganglionic spinal cord neurons (SPN), respectively. Careful timing of the survival of the animals allowed the detection of cell groups in immediate control of these DMV or SPN neurons. A far larger number of cell groups is involved in the control of DMV than of SPN neurons. Examples are given of a high level of interaction between the sympathetic and parasympathetic nervous system. Several cell groups project to both branches of the autonomic nervous system, sometimes even the same neurotransmitter is used, e.g., oxytocin neurons in the paraventricular nucleus and melanin-concentrating hormone and orexin neurons in the lateral hypothalamus project to both the DMV and SPN neurons. Moreover, the appearance of third-order neurons located in the sympathetic SPN after complete sympathectomy and in the DMV after complete vagotomy illustrates the possibility that motor neurons of the sympathetic and parasympathetic system may exchange information by means of interneurons. The presence of second-order neurons in prefrontal, gustatory, and piriform cortex may provide an anatomic basis for the involvement of these cortices in the cephalic insulin response. The observation that second-order neurons in both vagal and sympathetic control of the pancreas contain neuropeptides that are known to play a role in food intake indicates a direct association between behavioral and autonomic functions. Finally, the observation of third-order neurons in the suprachiasmatic nucleus and ventromedial hypothalamus shows the modulatory action of the time of the day and metabolic state, respectively.

Ghrelin is an appetite-stimulatory signal from stomach with structural resemblance to motilin.

BACKGROUND & AIMS: : Ghrelin, an endogenous ligand for growth hormone secretagogue receptor, was recently identified in the rat stomach. We examined the effects of the gastric peptide ghrelin on energy balance in association with leptin and vagal nerve activity. METHODS: : Food intake, oxygen consumption, gastric emptying, and hypothalamic neuropeptide Y (NPY) messenger RNA expression were measured after intra-third cerebroventricular or intraperitoneal injections of ghrelin in mice. The gastric vagal nerve activity was recorded after intravenous administration in rats. Gastric ghrelin gene expression was assessed by Northern blot analysis. Repeated coadministration of ghrelin and interleukin (IL)-1 beta was continued for 5 days. RESULTS: : Ghrelin exhibited gastroprokinetic activity with structural resemblance to motilin and potent orexigenic activity through action on the hypothalamic neuropeptide Y (NPY) and Y(1) receptor, which was lost after vagotomy. Ghrelin decreased gastric vagal afferent discharge in contrast to other anorexigenic peptides that increased the activity. Ghrelin gene expression in the stomach was increased by fasting and in ob/ob mice, and was decreased by administration of leptin and IL-1 beta. Peripherally administered ghrelin blocked IL-1 beta-induced anorexia and produced positive energy balance by promoting food intake and decreasing energy expenditure. CONCLUSIONS: : Ghrelin, which is negatively regulated by leptin and IL-1 beta, is secreted by the stomach and increases arcuate NPY expression, which in turn acts through Y(1) receptors to increase food intake and decrease energy expenditure. Gastric peptide ghrelin may thus function as part of the orexigenic pathway downstream from leptin and is a potential therapeutic target not only for obesity but also for anorexia and cachexia.

Leptin effects on feeding-related hypothalamic and peripheral neuronal activities in normal and obese rats.

We investigated the effects of leptin on central and/or peripheral feeding-related neuronal networks in Wistar male rats either normal (350-450 g) or Zucker obese (500-800 g). Low doses (1-10 pg) of leptin inhibited glucose-sensitive vagal hepatic afferent discharges and facilitated sympathetic efferent discharges to brown and white adipose tissue. Most (40-75%) neurons in the arcuate nucleus were significantly inhibited by superperfusion with leptin (0.1 nM-10 pM) under in vitro conditions. In anesthetized animals, leptin was applied electrophoretically to single hypothalamic neurons. Both glucose-sensitive neurons (GSNs) and non-GSNs in the feeding center (LHA) were significantly inhibited. Most glucoreceptor neurons in the satiety center (VMH) were significantly excited. Their depolarization was confirmed by activation of Na+ and K+ channels by 10(-11) M leptin using the perforate blind patch-clamp method. Although leptin excited GSNs in the parvocellular part of the paraventricular nucleus, the effects of leptin on such neuronal activity were slight or absent in Zucker obese rats. These results suggest that the feeding-suppression effects of leptin are mediated by its effects on signal transduction through both the central and the peripheral nervous systems.

Effect of infusion of vasoactive intestinal peptide (VIP)-antisense oligodeoxynucleotide into the third cerebral ventricle above the hypothalamic suprachiasmatic nucleus on the hyperglycemia caused by intracranial injection of 2-deoxy-D-glucose in rats.

We examined the effect of the infusion of a vasoactive intestinal peptide (VIP) antisense oligodeoxynucleotide into the third cerebral ventricle above the hypothalamic suprachiasmatic nucleus (SCN) using osmotic minipump for 3 days (0.2 nmol/ml per h) on the hyperglycemic response to intracerebroventricular injection of 2-deoxy-D-glucose (2DG) (80 micromol) in rats. After the infusion of the VIP antisense the inhibition of VIP expression in the SCN was observed in association with suppressions of the hyperglycemia, hyperglucagonemia and relative hypoinsulinemia due to the 2DG injection. Furthermore, additional intracranial injection of VIP (4 nmol) restored these responses to the 2DG injection in rats treated with the VIP antisense. These findings suggest that VIP neurons in the SCN are involved in the regulation of glucose metabolism.

Acidic fibroblast growth factor activates hypothalamic-pituitary-adrenocortical axis in rats.

Effects of acidic fibroblast growth factor (aFGF), an endogenous satiety substance, on the hypothalamic-pituitary-adrenocortical axis were examined under pentobarbital sodium anesthesia in rats. A guide cannula was inserted into the cerebral third ventricle and a vascular indwelling catheter was inserted into the right atrium from the jugular vein 2 wk and 3 days, respectively, before the experiment. A marked dose-dependent increase in plasma corticosterone was detected from 20 min to 2 h after intracerebroventricular administration of aFGF (1-10 ng). Significant increases in plasma adrenocorticotropic hormone (ACTH) were observed from 5 to 150 min after the intracerebroventricular administration of 10 ng aFGF. Significant dose-dependent increases in plasma corticosterone were also observed after intravenous injections of aFGF (1, 10, and 100 ng), together with increases in the plasma ACTH level. Pretreatment with antibody to corticotropin-releasing factor via the intracerebroventricular route abolished the increases in corticosterone induced by intracerebroventricularly administered aFGF, but not those induced by intravenous injection of aFGF. In adrenal glands perfused in situ with artificial medium, the corticosterone secretion rate increased slightly in response to 10(-9) M aFGF. These findings suggest that intracerebroventricular administration of aFGF activates the hypothalamic-pituitary-adrenal axis via corticotropin-releasing factor release in the brain, whereas peripheral administration of aFGF activates adrenocortical secretion mainly via a direct action on ACTH release.

Effect of oral administration of Pinellia ternata, Zingiberis rhizoma and their mixture on the efferent activity of the gastric branch of the vagus nerve in the rat.

The effect of taste stimulation of Pinellia temata, Zingiberis rhizoma and their mixture on the efferent activity of the gastric branch of the vagus nerve was observed in the anesthetized rat. Taste stimulation by Pinellia ternata (50 mg/ml, 10 min) resulted in a suppression in vagal gastric nerve activity. On the contrary, stimulation by Zingiberis rhizoma (50 mg/ml, 10 min) caused a facilitation in efferent activity. The mixture of Pinellia ternata and Zingiberis rhizoma (5:1, 50 mg/ml, 10 min stimulation) demonstrated no suppressive effect on gastric nerve activity. These observations indicate that it is reasonable to prescribe Pinellia ternata with Zingiberis rhizoma in traditional Japanese medicine to prevent suppressive effect of the taste of Pinellia ternata on gastric function.

Effect of byakko-ka-ninjin-to on the efferent activity of the autonomic nerve fibers innervating the sublingual gland of the rat.

The effect of intraduodenal infusion of Byakko-ka-ninjin-to (BN) on the efferent activity of the autonomic outflow to the sublingual gland was observed in the anesthetized rat. Intraduodenal infusion of BN (50-200 mg/kg) resulted in a dose-related increase in efferent activity. The enhancement of the nerve activity following administration of 200 mg/kg BN lasted longer than three hours. It was observed that the suppressive effect on efferent activity due to i.v. administration of hypertonic saline was antagonized by intraduodenal infusion of BN. From these observations it is suggested that Byakko-ka-ninjin-to acts as a facilitatory agent on salivary secretion.

Effects of Hoelen on the efferent activity of the gastric vagus nerve in the rat.

Hoelen is used to treat gastric disease in Eastern traditional medicine. The efferent activity on the gastric vagus nerve was studied in the rat, and the activity was found to increase after administration of Hoelen into the duodenum. In addition, we examined the effect of Hoelen fractions. Both the fraction containing polysaccharide and the triterpenoid-rich fraction increased activity significantly. Hoelen would thus appear useful for treating gastric disease by activating efferent activity of the gastric vagus nerve through the action of triterpenoid and polysaccharide.

An electrophysiological study on the vagal innervation of the thymus in the rat.

Vagal innervation of the thymus was studied by means of electrophysiological technique in the rat. Under urethane anesthesia, evoked action potentials originated from cervical vagus by electrical stimulations were recorded from the central cut end of the thymic branch of the vagus nerve after averaging for 32 times. It was observed that the conduction velocities are distributed in the range of 0.56-6.84 m/s, and the majority of vagal fibers in the thymic branch of the vagus nerve belong to a nonmyelinated C-fiber group. Further, it was confirmed that the right and left lobes of the thymus are innervated by cervical vagi bilaterally. The results suggest that the thymic branch of the vagus nerve plays a role in modulation of thymic function.

Parenteral nutrients in rat suppresses hepatic vagal afferent signals from portal vein to hypothalamus.

BACKGROUND: Parenteral nutrition (PN) suppresses spontaneous food intake in rats by stimulating dopamine in lateral hypothalamic area (LHA), which is attenuated by vagotomy. Sensors for glucose and for some individual amino acids exist in hepatoportal areas sending signals via hepatic vagal afferents to LHA. Hypothesizing that the decrease in spontaneous food intake occurs because a PN amino acid solution is recognized by portal vein sensors, we measured afferent nerve discharges in hepatic vagus in response to PN. METHODS: Hepatic vagus was divided in 11 rats, and an isolated nerve filament was placed on recording electrodes. Afferent firing rate was recorded before and after injecting during 1 minute a well-balanced crystalline amino acid mixture into either the portal or peripheral vein. Isotonic glucose was used as standard solution. RESULTS: A significant decrease in hepatic vagal afferent discharge rate was recorded after both intraportal and intravenous injection of the PN amino acid mixture. The mean discharge rate was inversely related to the concentration of the solution. Successive injections produced further decreases in discharge rate, and the effect was more sustained. CONCLUSIONS: The observed decrease in discharge rates suggests that the crystalline amino acid PN solution is sensed in hepatoportal areas informing LHA via hepatic afferents. This effect may play a role in regulating food intake during PN.

Effect of Pinellia ternata tuber on the efferent activity of the gastric vagus nerve in the rat.

Effect of intraduodenal infusion with the hot aqueous extract of Pinellia ternata tuber on the efferent discharges in the gastric branch of the vagus nerve was observed in the anesthetized rat. The infusion of the extract in doses of 2-15O mg per animal (c.a. 300 g, b.wt.) resulted in a dose-related increase in efferent activity of the vagal gastric nerve. The enhancement of the nerve activity following administration of 150 mg of this substance lasted longer than 90 min. It was observed that the suppressive effect on vagal gastric activity due to apomorphine and copper sulfate was antagonized by prior administration of the extract. From these observations it is suggested that Pinellia tuber acts as a facilitatory agent on gastric function.

Effects of hypothalamic stimulation and lesion on adrenal nerve activity.

Activity changes of efferent adrenal sympathetic nerves in response to bilateral manipulations of the hypothalamus, partly after intra-third cerebroventricular injection of 2-deoxy-D-glucose (2-DG) were investigated in anesthetized rats. Stimulation of the middle part of the lateral hypothalamic area (LHAm) increased adrenal nerve activity, whereas lesion caused rapid and remarkable decrease. Stimulation of the anterior part of the LHA (LHAa) tended to decrease the activity, and lesion produced either rapid decrease or late moderate increase. Stimulation of the ventromedial hypothalamic nucleus (VMH) did not affect the nerve activity, but lesion increased it gradually and then remarkably. Cerebroventricular infusion of 2-DG caused remarkable increase in activity that was suppressed by LHAm lesion. Subsequent infusion of 2-DG during the period of suppressed activity was no longer effective. The increased firing rate after 2-DG was suppressed by stimulation of the VMH, whereas lesion caused no change. These findings indicate that the central regulation of adrenal nerve activity is connected with individual hypothalamic regions and consequently depends on the degree and mode of activation of the sympathoadrenal system.

Neural control of blood glucose level.

All of the experimental results described above can be categorized as follows: the relationship between glucose levels and pancreatic and adrenal nerve activities; innervations of the liver and their role in the regulation of blood glucose level; central integration of blood glucose level; glucose-sensitive afferent nerve fibers in the liver and regulation of blood glucose; oral and intestinal inputs involved in reflex control of blood glucose level. We showed that an increase in blood glucose content produced an increase in the activity of the pancreatic branch of the vagus nerve, whereas it induced a decrease in the activity of the adrenal nerve. It was also shown that a decrease in blood glucose activated the sympatho-adrenal system and suppressed the vago-pancreatic system. It seems rational that these responses are involved in the maintenance of blood glucose level. Studies on the innervation of the liver led us to a conclusion that sympathetic innervation of the liver might play a role in eliciting a prompt hyperglycemic response through liberation of norepinephrine from the nerve terminals, and that the vagal innervation synergically worked with the humoral factor (insulin) for glycogen synthesis in the hyperglycemic condition. The glucose-sensitive afferents from the liver seem to initiate a reflex control of blood glucose level. The gustatory information on EIR response, reported by STEFFENS, is supported by the electrophysiological observations. MEI's reports also indicated the importance of information from the intestinal glucoreceptors in the reflex control of insulin secretion. The role of integrative functions of the hypothalamus and brainstem through neuronal networks on neural control of blood glucose levels is also evident. A schematic diagram of the nervous networks involved in the regulation of the blood glucose levels is shown in Fig. 3.

Effects of hypothalamic lesion on pancreatic autonomic nerve activity in the rat.

The effects of hypothalamic lesions and intravenous glucose infusion on the efferent activity of vagal and splanchnic nerves to the pancreas were studied in anesthetized rats. Lesions of the ventromedial hypothalamic (VMH), the dorsomedial hypothalamic (DMH) and the paraventricular (PVN) nuclei increased vagal and reduced splanchnic nerve activity. Lesion of the lateral hypothalamic area (LHA) decreased pancreatic vagal nerve activity, and produced either increased or decreased activity of pancreatic splanchnic nerve. Intravenous glucose infusion increased activity of the vagal nerve and reduced that of the splanchnic nerve. These glucose responses were influenced by hypothalamic lesions only slightly or not at all. The findings suggest that hypothalamic modulation of pancreatic hormone secretion involves both the parasympathetic and sympathetic nervous systems, and provide evidence that not only the VMH and the LHA but also the DMH and the PVN are involved in this mechanism.