Separation of hepatic and gastrointestinal signals from the common "hepatic" branch of the vagus

Anatomic studies show that the common hepatic branch (CHB) of the vagus contains afferent fibers that innervate sites outside the hepatoportal region, primarily in the gastrointestinal tract. In the current experiments on the anesthetized rat, the source of signals from the CHB was determined by recording CHB neurophysiological responses before and after transection of the gastroduodenal branch (GDB) of the CHB. Serotonin [5-hydroxytryptamine (5-HT)] and CCK-8 were used as probes to stimulate the CHB. Most of the CHB afferent fibers were 5-HT sensitive (56%), and 35% of these were also sensitive to CCK-8. Portal vein vs. jugular vein infusion of 5-HT and CCK-8 and GDB transection showed that 5-HT- and CCK-sensitive fibers innervate the hepatoportal region and areas outside the hepatic hilus (e.g., the gastrointestinal tract). Suppression of basal nerve activity by a 5-HT(3) receptor antagonist (Y-25130) suggests that approximately 50% of CHB afferent fibers contain 5-HT(3) receptors, but none of these fibers appears to be in the hepatoportal region because only in rats with an intact GDB did Y-25130 reduce nerve activity. In summary, these data are in close agreement with anatomic observations on the distribution of the CHB fibers and indicate that neurophysiological studies of the CHB must be carefully evaluated given the prominent role of nonhepatoportal afferent signals recorded from the CHB.     

The gastroduodenal branch of the common hepatic vagus regulates voluntary lard intake, fat deposition, and plasma metabolites in streptozotocin-diabetic rats

The common hepatic branch of the vagus nerve negatively regulates lard intake in rats with streptozotocin (STZ)-induced, insulin-dependent diabetes. However, this branch consists of two subbranches: the hepatic branch proper, which serves the liver, and the gastroduodenal branch, which serves the distal stomach, pancreas, and duodenum. The aim of this study was to determine whether the gastroduodenal branch specifically regulates voluntary lard intake. We performed a gastroduodenal branch vagotomy (GV) on nondiabetic, STZ-diabetic, and STZ-diabetic insulin-treated groups of rats and compared them with sham-operated counterparts. All rats had high steady-state corticosterone levels to maximize lard intake. Five days after surgery, all rats were provided with the choice of chow or lard to eat for another 5 days. STZ-diabetes resulted in a reduction in lard intake that was partially rescued by either GV or insulin treatment. Patterns of white adipose tissue (WAT) deposition differed after GV- and insulin-induced lard intake, with subcutaneous WAT increasing exclusively after the former and mesenteric WAT increasing exclusively in the latter. GV also prevented the insulin-induced reduction in the STZ-elevated plasma glucagon, triglycerides, free fatty acids, and total ketone bodies but did not alter the effect of insulin-induced reduction of plasma glucose levels. These data suggest that the gastroduodenal branch of the vagus inhibits lard intake and regulates WAT deposition and plasma metabolite levels in STZ-diabetic rats.     

Inhibitory effects on intake of cholecystokinin-8 and cholecystokinin-33 in rats with hepatic proper or common hepatic branch vagal innervation

The relative potencies of cholecystokinin (CCK)-8 and CCK-33 for decreasing meal size depend on the route of administration. Inhibitory potencies are equal after intraperitoneal administration, but CCK-33 is significantly more potent after intraportal administration. This suggests that CCK-33 is a more effective stimulant of hepatic afferent vagal nerves than is CCK-8. To investigate this possibility, we administered both peptides intraperitoneally in rats with abdominal vagotomies that spared only the hepatic proper vagal nerves (H) and in rats with abdominal vagotomies that spared the common hepatic branch that contains the fibers of the hepatic proper and gastroduodenal nerves (HGD). The vagal afferent innervation in H and HGD rats was verified with a wheat germ agglutinin-horseradish tracer strategy. Intraperitoneal administration of CCK-33 decreased 30-min intake of 10% sucrose in H rats as much as in sham rats, but CCK-8 decreased intake significantly less in H rats than in sham rats. The larger inhibitory effect of CCK-33 than of CCK-8 in H rats is consistent with the hypothesis that CCK-33 is a more effective stimulant of the hepatic proper vagal afferent nerves than CCK-8. In contrast to the results in H rats, the inhibitory potencies of both peptides were significantly and equivalently reduced in HGD rats compared with sham rats. This suggests that there is an inhibitory interaction between the stimulation of the gastroduodenal and hepatic proper afferent fibers by CCK-33.     

Peptides that regulate food intake: norepinephrine is not required for reduction of feeding induced by cholecystokinin

CCK octapeptide (CCK-8) is released by the gut in response to a meal and acts via CCK(A) receptors on vagal afferents to induce satiety. However, the central neural pathways by which peripheral CCK-8 affects feeding are poorly understood. In the present study, we tested the hypothesis that norepinephrine (NE) is necessary for satiety induced by peripheral CCK-8 by using mice lacking dopamine beta-hydroxylase (Dbh(-/-)), the enzyme responsible for synthesizing NE and epinephrine from dopamine. We found that Dbh(-/-) mice are as responsive to the satiating effects of CCK-8 as their normal littermates.     

Dectin-1 is not required for the host defense to Cryptococcus neoformans

Dectin-1 is known as a sole receptor for beta-glucan, a major cell wall component of fungal microorganisms. In the current study, we examined the role of this molecule in the host defense to Cryptococcus neoformans, an opportunistic fungal pathogen in AIDS patients. There was no significant difference in the clinical course and cytokine production between dectin-1 gene-deficient and control mice. These results indicate that dectin-1 is not likely essential for the development of host protective responses to C. neoformans.     

Endogenous cholecystokinin is not a major regulator of food intake in the chicken

This study investigated whether or not endogenous cholecystokinin exerts satiety effects in chickens. After several doses (0, 1, 2 and 4 g·kg body weight^-1) of intravenous injection of caerulein, the bile flow was increased in a dose-dependent fashion. However, the pharmacological level of caerulein failed to suppress the food intake of chickens. Two potent stimulators of endogenous cholecystokinin, i.e., soybean trypsin inhibitor and phenylalanine were administered to chickens before feeding and food intake was determined over 2 h. The soybean trypsin inhibitor and phenylalanine did not alter food intake. Devazepide, a cholecystokinin-A receptor antagonist, significantly decreased amylase release from the dispersed chicken pancreatic acini stimulated by caerulein. However, devazepide did not improve food intake of the chicken. The results obtained suggest that endogenous cholecystokinin may not act as a satiety signal in chickens.Abbreviations BSA bovine serum albumin - BW body weight - CCK cholecystokinin - DVZ devazepide - Hepes N-2-Hydroxyethylpiperazine-N-2-ethanesulfonic acid - i.p. intraperitoneal - i.v. intravenous - Phe phenylalanine - SBTI soybean trypsin inhibitor - SEM standard errors of means     

Exploring the glycemic response to food intake with undergraduate students at the University of La Reunion

Diabetes constitutes an increasingly prevalent disease, dramatically associated with an enhanced mortality risk in the developed world. A high prevalence of diabetes has recently been described at Réunion Island, a French department located in the Indian Ocean. At the University of La Réunion, a laboratory course involving students was designed to teach them blood glucose measurements and to examine the influence of food intake on their glycemic response. Using glucose meters, test strips, lancet devices, and sterile lancets, students determined their basal and postprandial glycemia. After plotting the variation over time of their glycemia, students calculated their glycemic response to a meal as the area under the curve. First, students observed that their glycemia had increased rapidly after food intake to values of <1.4 g/l and then decreased to normal values, proving the existence of a physiological regulatory system for glycemia. Using impedance balances, students then determined their body mass index and fat mass percentages. Positive and significant correlations were established between students' fat mass percentages and the glycemic response to the meal. A higher postprandial response was indeed noticed for students having higher fat percentages. Therefore, this laboratory allows students to observe the regulation of glycemia. It also alerts them to the correlation between higher body fat content and a higher glycemic response, which can be related to diabetic disorders. This laboratory constitutes an active illustration of their plenary lesson in endocrinology and particularly for the session dealing with glucose regulation.     

The individual and combined effects of glycemic index and protein on glycemic response, hunger, and energy intake

Although high protein and low glycemic index (GI) foods are thought to promote satiety, little is known about the effects of GI, protein, and their interaction on hunger and energy intake several hours following a mixed meal. This study investigated the long term effects of GI, protein, and their combined effects on glucose, insulin, hunger, and energy intake in healthy, sedentary, overweight, and obese adults (BMI of 30.9 ± 3.7 kg/m²). Sixteen individuals participated separately in four testing sessions after an overnight fast. The majority (75%) were non‐Hispanic Blacks. Each consumed one of four breakfast meals (high GI/low protein, high GI/high protein, low GI/low protein, low GI/high protein) in random order. Visual analog scales (VAS) and blood samples were taken at baseline, 15 min, and at 30 min intervals over 4 h following the meal. After 4 h, participants were given the opportunity to consume food ad libitum from a buffet style lunch. Meals containing low GI foods produced a smaller glucose (P < 0.002) and insulin (P = 0.0001) response than meals containing high GI foods. No main effects for protein or interactions between GI and protein were observed in glucose or insulin responses, respectively. The four meals had no differential effect on observed energy intake or self‐reported hunger, satiety, and prospective energy intake. Low GI meals produced the smallest postprandial increases in glucose and insulin. There were no effects for GI, protein, or their interaction on appetite or energy intake 4 h after breakfast.     

The repair and recombination enzyme ERCC1 is not required for immunoglobulin class switching

Class switch recombination (CSR) is a programmed gene rearrangement in which a B cell which is producing IgM and IgD antibody develops into an IgG-, IgA- or IgE-expressing cell. This is achieved by recombination between switch regions located 5' of each of the immunoglobulin heavy chain constant regions, except Cdelta. The mechanism of CSR has not been resolved but it is thought to involve a double-strand break followed by end joining. It has previously been suggested that the nucleotide excision repair protein ERCC1 may be involved in CSR due to its known roles in removal of 3' single-stranded tails in various types of recombination. In this study, we examined class switching in cultured splenocytes from ERCC1-deficient mice and found no evidence of any deficiency.     

Osteopontin is not required for the development of Th1 responses and viral immunity

Osteopontin (OPN) has been defined as a key cytokine promoting the release of IL-12 and hence inducing the development of protective cell-mediated immunity to viruses and intracellular pathogens. To further characterize the role of OPN in antiviral immunity, OPN-deficient (OPN-/-) mice were analyzed after infection with influenza virus and vaccinia virus. Surprisingly, we found that viral clearance, lung inflammation, and recruitment of effector T cells to the lung were unaffected in OPN-/- mice after influenza infection. Furthermore, effector status of T cells was normal as demonstrated by normal IFN-gamma production and CTL lytic activity. Moreover, activation and Th1 differentiation of naive TCR transgenic CD4+ T cells by dendritic cells and cognate Ag was normal in the absence of OPN in vitro. Contrary to a previous report, we found that OPN-/- mice mounted a normal immune response to Listeria monocytogenes. In conclusion, OPN is dispensable for antiviral immune responses against influenza virus and vaccinia virus.     

The germ cell determinant Blimp1 is not required for derivation of pluripotent stem cells

Blimp1 (Prdm1), the key determinant of primordial germ cells (PGCs), plays a combinatorial role with Prdm14 during PGC specification from postimplantation epiblast cells. They together initiate epigenetic reprogramming in early germ cells toward an underlying pluripotent state, which is equivalent to embryonic stem cells (ESCs). Whereas Prdm14 alone can?promote reprogramming and is important for the propagation of the pluripotent state, it is not known whether Blimp1 is similarly involved. By using a genetic approach, we demonstrate that Blimp1 is?dispensable for the derivation and maintenance of ESCs and postimplantation epiblast stem cells (epiSCs). Notably, Blimp1 is also dispensable for reprogramming epiSCs to ESCs. Thus, although Blimp1 is obligatory for PGC specification, it is not required for the reversion of epiSCs to ESCs and for their maintenance thereafter. This study suggests that reprogramming, including that of somatic cells to ESCs, may not entail an obligatory route through a Blimp1-positive PGC-like state.     

The major subunit of Escherichia coli type 1 fimbriae is not required for D-mannose-specific adhesion

Type 1 fimbriae are surface organelles on Escherichia coli, which mediate specific binding to D-mannose-containing structures. These fimbriae are heteropolymers composed of a major building element, the FimA protein, and small amounts of the FimF, FimG and FimH proteins. The FimH protein is uniquely responsible for the D-mannose receptor binding. In this work data are presented which indicate that the major subunit of type 1 fimbriae is dispensable for D-mannose-specific binding. A recombinant strain was studied which harboured an insertional deletion in the fimA gene, and was thereby unable to produce type 1 fimbriae; however, it was still able to express a D-mannose-binding phenotype. However, the deletion resulted in a 25-fold reduction of the adhesive potential, as measured by binding to D-mannose-coated Sepharose beads. Serological and specific receptor binding evidence is presented that suggests that the FimH adhesion is capable of being exposed on the bacterial surface without being an integral part of the fimbriae.     

The tandem BRCT domain of 53BP1 is not required for its repair function

53BP1 plays an important role in cellular response to DNA damage. It is thought to be the mammalian homologue of budding yeast Rad9 and/or fission yeast Crb2. Rad9/Crb2 are bona fide checkpoint proteins whose activation requires their corresponding C-terminal tandem BRCT (BRCA1 C-terminal) motifs, which mediate their oligomerization and phosphorylation at multiple sites following DNA damage. Here we show that the function of human 53BP1 similarly depends on its oligomerization and phosphorylation at multiple sites but in a BRCT domain-independent manner. Moreover, unlike its proposed yeast counterparts, human 53BP1 only has limited checkpoint functions but rather acts as an adaptor in the repair of DNA double strand breaks. This difference in function may reflect the higher complexity of the DNA damage response network in metazoa including the evolution of other BRCT domain-containing proteins that may have functions redundant or overlapping with those of 53BP1.     

Y1 and Y5 receptors are both required for the regulation of food intake and energy homeostasis in mice

Neuropeptide Y (NPY) acting in the hypothalamus is one of the most powerful orexigenic agents known. Of the five known Y receptors, hypothalamic Y1 and Y5 have been most strongly implicated in mediating hyperphagic effects. However, knockout of individual Y1 or Y5 receptors induces late-onset obesity--and Y5 receptor knockout also induces hyperphagia, possibly due to redundancy in functions of these genes. Here we show that food intake in mice requires the combined actions of both Y1 and Y5 receptors. Germline Y1Y5 ablation in Y1Y5(-/-) mice results in hypophagia, an effect that is at least partially mediated by the hypothalamus, since mice with adult-onset Y1Y5 receptor dual ablation targeted to the paraventricular nucleus (PVN) of the hypothalamus (Y1Y5(Hyp/Hyp)) also exhibit reduced spontaneous or fasting-induced food intake when fed a high fat diet. Interestingly, despite hypophagia, mice with germline or hypothalamus-specific Y1Y5 deficiency exhibited increased body weight and/or increased adiposity, possibly due to compensatory responses to gene deletion, such as the decreased energy expenditure observed in male Y1Y5(-/-) animals relative to wildtype values. While Y1 and Y5 receptors expressed in other hypothalamic areas besides the PVN--such as the dorsomedial nucleus and the ventromedial hypothalamus--cannot be excluded from having a role in the regulation of food intake, these studies demonstrate the pivotal, combined role of both Y1 and Y5 receptors in the mediation of food intake.     

The presequence of yeast 5-aminolevulinate synthase is not required for targeting to mitochondria

A truncated form of the yeast mitochondrial 5-aminolevulinate (ALA) synthase was constructed by deletion of the first 75 amino acid residues of its precursor form. This truncated ALA synthase which lost its entire presequence and 40 residues of the mature part possesses a new amino terminus quite different from a typical mitochondrial presequence. This modified protein expressed in vivo is found entirely located within mitochondria. Although it was now unable to reach the matrix space, it was internalized as shown by its resistance to protease in isolated mitochondria. Pulse-chase radiolabeling in the presence of an uncoupler suggests that a membrane potential is not required for the targeting of this truncated ALA synthase. Thus, the amino-terminal signal, if indispensable as a matrix targeting signal, could be replaced by an internal sequence or a particular folding for recognition by the import machinery.     

The absence of Ku but not defects in classical non-homologous end-joining is required to trigger PARP1-dependent end-joining

Classical-non-homologous end-joining (C-NHEJ) is considered the main pathway for repairing DNA double strand breaks (DSB) in mammalian cells. When C-NHEJ is defective, cells may switch DSB repair to an alternative-end-joining, which depends on PARP1 and is more erroneous. This PARP1-EJ is suggested to be active especially in tumor cells contributing to their genomic instability. Here, we define conditions under which cells would switch the repair to PARP1-EJ. Using the end jining repair substrate pEJ, we revealed that PARP1-EJ is solely used when Ku is deficient but not when either DNA-PKcs or Xrcc4 is lacking. In the latter case, DSB repair, however, could be shuttled to PARP1-EJ after additional Ku80 down-regulation, which partly rescued the DSB repair in these mutants. We demonstrate here that PARP-EJ may work on DSB ends at high fidelity manner, as evident from the unchanged efficiency upon blocking end resection by either roscovitin or mirin. Furthermore, we demonstrate for that PARP-EJ is likewise involved in the repair of multiple DSBs (I-PpoI- and IR-induced). Importantly, we identified a chromatin signature associated with the switch to PARP1-EJ which is characterized by a strong enrichment of both PARP1 and LigIII at damaged chromatin. Together, these data indicate that Ku is the main regulator for the hierarchal organization between C-NHEJ and PARP1-EJ.     

Adaptation is not required to explain the long-term response of axons to molecular gradients

It has been suggested that growth cones navigating through the developing nervous system might display adaptation, so that their response to gradient signals is conserved over wide variations in ligand concentration. Recently however, a new chemotaxis assay that allows the effect of gradient parameters on axonal trajectories to be finely varied has revealed a decline in gradient sensitivity on either side of an optimal concentration. We show that this behavior can be quantitatively reproduced with a computational model of axonal chemotaxis that does not employ explicit adaptation. Two crucial components of this model required to reproduce the observed sensitivity are spatial and temporal averaging. These can be interpreted as corresponding, respectively, to the spatial spread of signaling effects downstream from receptor binding, and to the finite time over which these signaling effects decay. For spatial averaging, the model predicts that an effective range of roughly one-third of the extent of the growth cone is optimal for detecting small gradient signals. For temporal decay, a timescale of about 3 minutes is required for the model to reproduce the experimentally observed sensitivity.