Autonomic Neuroscience: Basic and Clinical 85 (2000) 98–101 www.elsevier.com / locate / autneu
Short communication
Antagonism of central glucagon-like peptide-1 receptors enhances lipopolysaccharide-induced fever Linda Rinaman*, Jason Comer
1
Department of Neuroscience, 446 Crawford Hall, University of Pittsburgh, Pittsburgh, PA 15260, USA
Abstract Lipopolysaccharide (LPS; a model of systemic bacterial infection) causes fever and activates glucagon-like peptide-1 (GLP-1) neurons in the caudal brainstem. The present study examined whether central GLP-1 receptor signaling plays a functional role in LPS-induced fever. Adult male Sprague–Dawley rats were injected i.p. with LPS (0 or 100 mg / kg), then were infused intracerebroventricularly with GLP-1 receptor antagonist (0 or 10 mg) delivered 2.5 h after injection of LPS or vehicle. Core body temperature was measured at 30-min intervals for 6.5 h after LPS treatment. Consistent with previous reports, body temperature was significantly elevated within 90 min and remained elevated for the remainder of the monitoring period. The pyrogenic effect of LPS was amplified in rats that received central infusion of GLP-1 receptor antagonist, although the antagonist by itself did not alter body temperature. These findings suggest that endogenous GLP-1 acts at central receptors to limit the fever response in rats after i.p. administration of LPS. 2000 Elsevier Science B.V. All rights reserved. Keywords: Thermoregulation; Pyrogen; Vagus
1. Introduction Systemic administration of lipopolysaccharide (LPS), the protein-free form of bacterial endotoxin, causes fever in rats and other mammals (see Blatteis and Sehic, 1998). A recent study showed that intraperitoneal (i.p.) administration of a pyrogenic dose of LPS (100 mg / kg) in rats activates expression of the immediate-early gene product, cFos, in hindbrain neurons that are immunoreactive for glucagon-like peptide-1 (GLP-1) (Rinaman, 1999b). A possible thermoregulatory role for GLP-1 has been proposed based on evidence that intracerebroventricular (i.c.v.) or i.p. administration of synthetic GLP-1 reduces baseline body (core) temperature (T c ) in rats (O’Shea et al., 1996). GLP-1-positive neurons are located in the medullary dorsal vagal complex and adjacent reticular formation (Alvarez et al., 1996; Larsen et al., 1997), and thus are ideally positioned to receive direct or relayed neural signals from subdiaphragmatic vagal afferents and other *Corresponding author. Tel.: 11-412-624-6994; fax: 11-412-6249198. E-mail address:
[email protected] (L. Rinaman). 1 Present address: Graduate Program, Department of Biology, Duquesne University, Pittsburgh, PA 15282, USA.
central neural components of the fever response to i.p. LPS (see Elmquist et al., 1997; Romanovsky et al., 1997). The present study was designed to test the hypothesis that endogenous GLP-1 acting at central receptors participates in the pyrogenic effect of systemically administered LPS.
2. Methods Experimental procedures were reviewed and approved by the University of Pittsburgh Institutional Animal Care and Use Committee. Male Sprague–Dawley rats (Zivic Miller; Zelionople, PA, USA; 225–275 g) were housed individually in hanging wire cages in a controlled environment (248C, lights on from 0700 to 1900 h) with free access to water and pelleted rat chow (Purina). Rats were anesthetized and fitted with chronic indwelling 26-gauge stainless steel guide cannulas (Plastics One) aimed at the lateral ventricle as described previously (Rinaman, 1999a). Correct cannula placement was verified 1 week after surgery by measuring the drinking response to i.c.v. angiotensin II in water-replete rats, as previously described (Rinaman, 1999a). All rats used in the present study (n516) drank at least 8.0 ml of water within 30 min of i.c.v. administration of 10 ng angiotensin II (Sigma).
1566-0702 / 00 / $ – see front matter 2000 Elsevier Science B.V. All rights reserved. PII: S1566-0702( 00 )00227-7
L. Rinaman, J. Comer / Autonomic Neuroscience: Basic and Clinical 85 (2000) 98 – 101
The T c was measured by using an electronic digital thermometer with a small flexible rectal probe (Traceable; VWR Scientific) inserted 5 cm into the colon. Rats were acclimated to this procedure by having T c measured three times a day for 2 days before the experiment. On the day of the experiment, baseline T c was determined at 1000 h, then rats were immediately injected i.p. with LPS (100 mg / kg in 2.0 ml of 0.15 M NaCl; n516). T c was obtained from each rat at 30-min intervals for 6.5 h after LPS administration. After the 2.5 h post-LPS T c reading, when fever was well-established (see Results), rats were infused i.c.v. with 10 ml of sterile 0.15 M NaCl containing 0 or 10 mg of GLP-1 receptor antagonist (des His 1 Glu 9 -exendin 4; n58 rats per group). We and others (Thiele et al., 1998) have found that 10 mg of des His 1 Glu 9 -exendin 4 (American Peptides) administered i.c.v. in rats completely eliminates the potent anorexigenic effect of 10 mg of synthetic GLP-1 (7-36) amide infused i.c.v. (data not shown). An additional control experiment was performed 1 week later, using the same rats (n516) and a similar injection paradigm. After measuring baseline T c at 1000 h, rats were injected i.p. with 0.15 M NaCl (2.0 ml), followed 2.5 h later by i.c.v. infusion of 10 mg of the GLP-1 receptor antagonist (n516). T c was obtained from each rat at 30-min intervals for 6.5 h after the i.p. injection. T c values at each timepoint were combined by treatment group and are expressed as mean6S.E. Treatment-related differences in T c were tested for statistical significance by using two-way ANOVA. When F values indicated significant overall main treatment effects and interactions, the ANOVA was followed up with planned post hoc t-tests using Dunn’s (Bonferonni) correction for repeated measures analysis. Differences were considered significant when P,0.05.
3. Results Consistent with previous reports documenting the effects of LPS after i.p. administration, T c was significantly elevated above baseline within 90 min and remained elevated for the remainder of the monitoring period (Fig. 1). As shown in Fig. 1, central administration of des His 1 Glu 9 -exendin 4 (the GLP-1 receptor blocker) significantly elevated the LPS-induced increase in T c . The fever-enhancing effect of the GLP-1 receptor blocker was not statistically significant until 90 min after its infusion, although the effect persisted for the remainder of the T c monitoring period. Thus, pharmacological blockade of central GLP-1 receptors had a delayed-onset but persistent effect in exacerbating LPS-induced fever. In the absence of LPS treatment, however, central infusion of GLP-1 receptor antagonist produced no change in baseline T c (Fig. 1).
99
4. Discussion The results of this study suggest that endogenous central GLP-1 neural pathways play a significant functional role in limiting the fever produced by i.p. administration of LPS in rats. A potential role for GLP-1 in thermoregulation was previously inferred from the demonstration that synthetic GLP-1 (7–36) amide delivered i.c.v. significantly decreased baseline T c in rats, and that this effect could be blocked with a GLP-1 receptor antagonist (O’Shea et al., 1996). Our demonstration that central GLP-1 receptor blockade enhances the pyrogenic response to LPS treatment is consistent with those findings. In the absence of LPS-induced fever, baseline T c was not altered by central blockade of GLP-1 receptors in this or a previous study (O’Shea et al., 1996), evidence that endogenous central GLP-1 receptor signaling probably does not play a significant role in maintaining normal body temperature. Instead, treatment-induced activation of central GLP-1 neural pathways may provide a mechanism to limit the magnitude of the fever response. The precise location of GLP-1 neural pathways and receptors that might participate in controlling body temperature during fever remains unclear. Fever is a centrallycoordinated response that includes both endocrine and autonomic components (Elmquist et al., 1997) and central administration of synthetic GLP-1 produces marked effects on these general components of homeostatic control (see Blazquez et al., 1999). Increased central or vagal afferent signaling to the caudal medulla after LPS treatment could lead to the recruitment of hindbrain GLP-1 neurons, which are located in the dorsal vagal complex and adjacent reticular formation (Alvarez et al., 1996; Larsen et al., 1997). Supportive evidence for a vagally-mediated activation of GLP-1 neurons after LPS treatment comes from the finding that GLP-1 neurons are activated in rats after systemic administration of cholecystokinin (CCK) octapeptide (Rinaman, 1999b), which binds to receptors on subdiaphragmatic vagal afferents to increase vagal afferent signaling to the dorsal vagal complex (Schwartz et al., 1994). GLP-1-immunoreactive fibers and GLP-1 receptors have been localized in several brain regions implicated in body temperature regulation (Saper, 1998), including the ventromedial preoptic area, paraventricular nucleus of the hypothalamus, and nucleus of the solitary tract (Jin et al., 1988; Alvarez et al., 1996; Navarro et al., 1996; Larsen et al., 1997; Merchenthaler et al., 1999). Because GLP-1 receptor antagonist was infused into the lateral ventricle in the present study, the drug presumably had access to all of these central locations, any of which could potentially mediate the observed exacerbation of LPS-induced fever. It will be important to determine where endogenously released GLP-1 may be acting in the brain to control fever after LPS treatment, and whether central GLP-1 receptor signaling plays a role in other experimental and physiolog-
100
L. Rinaman, J. Comer / Autonomic Neuroscience: Basic and Clinical 85 (2000) 98 – 101
Fig. 1. Treatment-related changes in core body temperature (T c ) relative to baseline are plotted over time. Values are group means6S.E. The long horizontal bar (———) indicates timepoints at which T c is significantly increased above baseline in both LPS-treated groups (P,0.05 at each timepoint for each group). The short horizontal bar (—) indicates timepoints at which the increase in T c in rats that received LPS i.p. and the GLP-1 receptor blocker i.c.v. is significantly greater than in rats that received LPS i.p. and vehicle (NaCl) i.c.v. (P,0.05 at each timepoint). Rats received i.p. injections of LPS (100 mg / kg) or vehicle (0.15 M NaCl) immediately after the first baseline T c reading (0 min). Rats received i.c.v. infusion of GLP-1 receptor blocker (des His 1 Glu 9 -exendin 4; 10 mg) or vehicle (0.15 M NaCl) immediately after the 150 min T c reading, as indicated by the arrow.
ical circumstances that are associated with changes in body temperature.
Acknowledgements Jen-Shew Yen and Larry R. Murphy provided excellent technical assistance throughout the course of this study; supported by NIH grant No. MH59911.
References Alvarez, E., Roncero, I., Chowen, J.A., Thorens, B., Blazquez, E., 1996. Expression of the glucagon-like peptide-1 receptor gene in rat brain. J. Neurochem. 66, 920–927. Blatteis, C.M., Sehic, E., 1998. Cytokines and fever. Ann. NY Acad. Sci. 840, 608–618. Blazquez, E., Alvarez, E., Navarro, M., Roncero, I., Rodriguez-Fonseca, F., Chowen, J.A., Zueco, J.A., 1999. Glucagon-like peptide-1 (7–36) amide as a novel neuropeptide. Molec. Neurobiol. 18, 157–173.
Elmquist, J.K., Scammell, T.E., Saper, C.B., 1997. Mechanisms of CNS response to immune challenge: the febrile response. Trends Neurosci. 20, 565–570. Jin, S.L., Han, V.K., Simmons, J.G., Towle, A.C., Lauder, J.M., Lund, P.K., 1988. Distribution of glucagon-like peptide I (GLP-I), glucagon, and glicentin in the rat brain: an immunocytochemical study. J. Comp. Neurol. 271, 519–532. Larsen, P.J., Tang-Christensen, M., Holst, J.J., Orskov, C., 1997. Distribution of glucagon-like peptide-1 and other preproglucagon-derived peptides in rat hypothalamus and brainstem. Neuroscience 77, 257– 270. Merchenthaler, I., Lane, M., Shughrue, P., 1999. Distribution of pre-proglucagon and glucagon-like peptide-1 receptor messenger RNAs in the rat central nervous system. J. Comp. Neurol. 403, 261–280. Navarro, M., Fonseca, F.Rd., Alvarez, E., Chowen, J.A., Zueco, J.A., Gomez, R., Eng, J., Blazquez, E., 1996. Colocalization of glucagonlike peptide-1 (GLP-1) receptors, glucose transporter GLUT-2, and glucokinase mRNAs in rat hypothalamic cells: evidence for a role of GLP-1 receptor agonists as an inhibitory signal for food and water intake. J. Neurochem. 67, 1982–1991. O’Shea, D., Gunn, I., Chen, X., Bloom, S., Herbert, J., 1996. A role for central glucagon-like peptide-1 in temperature regulation. Neuroreport 7, 830–832. Rinaman, L., 1999a. A functional role for central glucagon-like peptide-1
L. Rinaman, J. Comer / Autonomic Neuroscience: Basic and Clinical 85 (2000) 98 – 101 receptors in lithium chloride-induced anorexia. Am. J. Physiol. 277, R1537–R1540. Rinaman, L., 1999b. Interoceptive stress activates glucagon-like peptide-1 neurons that project to the hypothalamus. Am. J. Physiol. 277, R582–R590. Romanovsky, A.A., Simons, C.T., Szekely, M., Kulchitsky, V.A., 1997. The vagus nerve in the thermoregulatory response to systemic inflammation. Am. J. Physiol. 272, R407–R413. Saper, C.B., 1998. Neurobiological basis of fever. Ann. NY Acad. Sci. 856, 90–94.
101
Schwartz, G.J., McHugh, P.R., Moran, T.H., 1994. Pharmacological dissociation of responses to CCK and gastric loads in rat mechanosensitive vagal afferents. Am. J. Physiol. 267, R303–R308. Thiele, T.E., Seeley, R.J., D’Alessio, D., Eng, J., Bernstein, I.L., Woods, S.C., van Dijk, G., 1998. Central infusion of glucagon-like peptide-1(7–36) amide (GLP-1) receptor antagonist attenuates lithium chloride-induced c-Fos induction in rat brainstem. Brain Res. 801, 164– 170.
