Fever and lethargy induced by subcutaneous pyrogen infusion in unrestrained rats

We have investigated the effects of continuous subcutaneous infusion of lipopolysaccharide (LPS), muramyldipeptide (MDP), or saline on abdominal temperature and voluntary activity in unrestrained rats. Both pyrogens were infused via osmotic pumps at a rate of approximately 2 microg.kg-1.min-1 for 7 d. LPS infusion evoked a 3-d and MDP a 1-d elevation in body temperature. Night-time activity was suppressed on days 1 and 2 during LPS infusion and on day 1 of MDP infusion. Body mass was significantly decreased on infusion day 4 in rats receiving either LPS or MDP; however, the rate of weight gain had been restored by day 8 (1 d after cessation of pyrogen infusion). We further tested the body temperature response of the same experimental animals to a single subcutaneous bolus injection (250 microg/kg) of the same pyrogen that had been infused for 7 d, 2 d after cessation of pyrogen infusion (day 9). The fever response in rats receiving a bolus injection of either LPS or MDP was significantly attenuated in rats that had previously been infused with the same pyrogen. These data suggest that tolerance developed to continuous infusion of both Gram-negative and Gram-positive pyrogens, and that mechanisms of tolerance development set in early during the 7-d infusion period of both pyrogens and persisted for at least 2 d after the cessation of pyrogen infusion. We propose that cytokine intermediates were involved or required in inducing these responses to continuous infusion of both LPS and MDP.     

Plasma concentration of prolactin during four-day osmotic pump infusion of thyrotropin-releasing hormone and vasoactive intestinal polypeptide in rats

Pituitary prolactin (PRL) responses to 4-day continuous infusion of thyrotropin-releasing hormone (TRH) and vasoactive intestinal polypeptide (VIP) were investigated in unanesthetized male rats using Alzet osmotic minipumps. The TRH dose infused was 3.6 micrograms/day and the VIP dose was 32.8 micrograms/day. Infusion of TRH with osmotic pumps elevated the plasma PRL level compared to controls over the 4-day infusion period. However, mean levels of PRL tended to decrease during the 4-day infusion. On the other hand, continuous VIP infusion elicited a significant continuous PRL release over the 4-day infusion period. Thus, it may be said that the PRL responses to infused TRH and VIP were maintained during the 4-day infusion.     

Effects of nitric oxide synthase inhibitors on the febrile response to muramyl dipeptide and lipopolysaccharide in rats

We have administered aminoguanidine, a relatively specific inhibitor of inducible nitric oxide synthase, and N-nitro-L-arginine methyl ester (L-NAME), an unspecific nitric oxide synthase inhibitor, to rats made febrile with the gram-positive pyrogen, muramyl dipeptide and gram-negative pyrogen, lipopolysaccharide. Sprague-Dawley rats, housed individually at approximately 25 degrees C with a 12:12 h light:dark cycle (lights on 0700 hours), were injected (at 0900 hours) intraperitoneally with 50 mg/kg aminoguanidine, 25 mg/kg or 50 mg/kg L-NAME, and intramuscularly with 500 microg/kg muramyl dipeptide or 100 microg/kg lipopolysaccharide. Pyrogen injections were spaced at least 14 days apart. Body temperature was measured throughout the study in unrestrained animals using radio-telemetry. Neither muramyl dipeptide nor lipopolysaccharide-induced fevers were affected by aminoguanidine. However, L-NAME administration inhibited muramyl dipeptide and lipopolysaccharide-induced fevers, but only for the 1st 2-4 h of the fevers (two-way ANOVA, P<0.05). After the initial inhibition, lipopolysaccharide fevers developed normally. Therefore, constitutively expressed nitric oxide synthase appears to be involved in the initial phases of fever genesis of gram-negative and gram-positive fevers in rats. On the other hand, inducible nitric oxide synthase appears not to play a role in these fevers.     

CCK-8 induces fever-like regulated hyperthermia and symptoms of sickness behavior in mice: A biotelemetric study

In earlier studies it has been found that rats respond to intracerebroventricular (i.c.v.) injection of cholecystokinin-octapeptide (CCK-8) with a febrile response characterized by rises of heat production and core temperature together with tail-skin vasoconstriction mediated by CCK2 receptors. Biotelemetric investigations of the same species have additionally shown that CCK-induced fever is accompanied by decreased locomotor activity. Similar data for mice have not been reported so far. In the present studies C57BL/6 mice were infused i.c.v. for 3 days with CCK-8 to see effects on body core temperature, locomotor activity, food intake and body weight. Biotelemetric monitoring disclosed a rise in daylight core temperature and a fall of night-time locomotor activity both lasting beyond the time of i.c.v. infusions. Food intake was suppressed only during infusion, while a significant decrease of body weight was sustained after the end of CCK-8 infusion. It is concluded that similar to rats mice also respond to i.c.v. infusion of CCK-8 with a fever-like (regulated) hyperthermia and some components of sickness behavior as measured by biotelemetry, and thus a CCK-mediated mechanism may contribute to fever genesis also in mice.     

Induction of premature delivery in sheep following infusion of cortisol to the fetus. I. The effect of maternal administration of progestagens

Premature induction of delivery in fetuses infused with graded doses of cortisol was brought about in 123.5 +/- 7.7 h (mean +/- SEM, n = 6) after the start of cortisol infusion. This treatment caused a rise in fetal plasma cortisol similar to that observed at normal delivery. Maternal and fetal progesterone and 20 alpha-dihydroprogesterone concentrations decreased to basal levels during infusion of cortisol to the fetus. Induction of premature delivery was delayed or prevented by concomitant treatment of the ewe with progestagen. Maternal intramuscular injection of 100 mg progesterone, 2 times daily, prevented delivery in four of four ewes treated during the time that cortisol was infused into the fetus (11-13 days). Maternal plasma progesterone and 20 alpha-dihydroprogesterone concentrations were maintained during this period, but fetal plasma progesterone concentrations decreased to the same extent as in the fetuses infused with cortisol alone. A single intramuscular injection of 250 mg of medroxyprogesterone acetate to ewes on the day before commencement of infusion of cortisol to the fetus prevented delivery in four of six ewes during the time that cortisol was infused for 9, 13, 14, and 15 days, respectively. One ewe delivered a live lamb at 133.5 h and another at 147.7 h after the start of infusion of cortisol to the fetus. Maternal and fetal plasma cortisol, progesterone, and 20 alpha-dihydroprogesterone concentrations were similar to those observed during infusion of cortisol alone to the fetus. Although fetal cortisol concentrations rose in a similar fashion, and to a similar extent, in all three groups during infusion of cortisol to the fetus, fetal 11-desoxycortisol concentrations only rose above basal levels close to the time of delivery in cortisol-infused fetuses or, in the progestagen-treated groups, when the fetus showed signs of being stressed.     

Hormone and haemodynamic effects of angiotensin II infusion during captopril treatment for heart failure

Withdrawal of captopril therapy for cardiac failure results in increments in plasma cortisol, noradrenaline and heart rate. To determine whether these changes related to the concomitant rise in circulating angiotensin II, we infused angiotensin II at 0.5, 2, 4 and 8 ng/kg/minute, each infusion lasting for 1 hour, in 4 patients during maintenance captopril therapy for heart failure. A control solution of 5% dextrose was infused over a similar time interval on a separate day. The study was performed under metabolic balance conditions, with constant body posture and continuous haemodynamic monitoring. Angiotensin II induced the expected rise in arterial pressure and in plasma aldosterone. In contrast the diurnal decline in plasma ACTH and cortisol was not altered, and no changes in noradrenaline or heart rate were observed. Plasma angiotensin II appears to have little or no effect on ACTH, cortisol, noradrenaline and heart rate under the conditions of this study.     

The effect of lipopolysaccharide on cholecystokinin in murine plasma and tissue

Several mechanisms have been proposed for neuroimmune communication supporting sickness behavior (fever, anorexia, inactivity, and cachexia) following infection. We examined the role of cholecystokinin as a neurochemical intermediary of sickness behavior by determining plasma, duodenum, hypothalamus, and brainstem cholecystokinin concentrations 30 and 60 min and 12 h following intraperitoneal lipopolysaccharide (LPS) (0.25 and 2.5 mg/kg). Hypothalamic cholecystokinin was significantly lower in LPS- versus saline-treated mice 30 min (0.25 and 2.5 mg/kg) and 12 h (2.5 mg/kg) post-injection. Plasma cholecystokinin of LPS-treated mice was significantly lower than that of controls 1 and 12 h post-injection, a finding consistent with a non-endocrine action of peripheral cholecystokinin.     

Effects of leptin on fetal plasma adrenocorticotropic hormone and cortisol concentrations and the timing of parturition in the sheep

We investigated whether leptin can suppress the prepartum activation of the fetal hypothalamus-pituitary-adrenal (HPA) axis and delay the timing of parturition in the sheep. First, we investigated the effects of a 4-day intravascular infusion of recombinant ovine leptin (n = 7) or saline (n = 6) on fetal plasma adrenocorticotropic hormone (ACTH) and cortisol concentrations, starting from 136 days gestation (i.e., at the onset of the prepartum activation of the fetal HPA axis. The effects of a continuous intrafetal infusion of leptin (n = 7) or saline (n = 5) from 144 days gestation on fetal plasma ACTH and cortisol concentrations and the timing of delivery were also determined in a separate study. There was an increase in fetal plasma ACTH (P < 0.01) and cortisol (P < 0.001) concentrations when saline was infused between 136-137 and 140-141 days gestation. Plasma ACTH and cortisol concentrations did not rise, however, when leptin was infused during this period of gestation. When leptin was infused after 144 days gestation, there was no effect of a 4- to 5-fold increase in circulating leptin on fetal ACTH concentrations. In contrast, leptin infusion from 144 days gestation suppressed (P < 0.05) fetal plasma cortisol concentrations by around 40% between 90 and 42 h before delivery. There was no difference, however, in the length of gestation between the saline- and leptin-infused groups (saline infused, 150.2 +/- 0.5 days; leptin infused, 149.8 +/- 1.0 days). In saline-infused fetuses, there was a significant negative relationship between the plasma concentrations of cortisol (y) and leptin (x) between 138 and 146 days gestation (y = 81.4 - 7.7x, r = 0.38, P < 0.005). This study provides evidence for an endocrine negative feedback loop between leptin and the HPA axis in fetal life.     

Continuous central vasopressin infusion increases peripheral fetal corticotropin and cortisol in the fetal sheep

In previous studies on regulation of fetal adrenocorticotropin (ACTH) secretion, corticotropin releasing factor (CRF) and arginine vasopressin (AVP) have been administered by peripheral intravascular infusion. In order to look at an alternate route of administration, we investigated the effect of continuous intracerebroventricular administration of AVP to the fetus on fetal plasma ACTH and fetal and maternal plasma cortisol concentrations. Sheep fetuses (n = 9) were instrumental with carotid artery and lateral cerebral ventricular catheters. Fetuses were given intracerebroventricular infusion from 125-134 days gestational age of artificial cerebrospinal fluid vehicle (n = 4), or AVP 250 mu U.min-1 continuously in artificial cerebrospinal fluid vehicle (n =5). Fetal blood was obtained daily between 09.00 and 12.00h and 20.00 and 23.00h. Over the infusion period, fetal plasma ACTH and cortisol concentrations in AVP infused fetuses increased (P less than 0.05) compared with the vehicle infused group. Gestation length for the fetuses in the AVP and vehicle infused groups were 139 +/- 4.9 (n =4) and 145 +/- 4.6 (n = 3) days respectively (n.s.). Fetal plasma AVP concentrations in the AVP infused group were not different from the vehicle infused group.     

Influence of oxytocin infusion during oestrus and the early luteal phase on progesterone secretion and the establishment of pregnancy in ewes

In Experiment 1, an osmotic minipump containing oxytocin was implanted s.c. in ewes for 12 days beginning on Day 10 of the oestrous cycle, producing approximately 100 pg oxytocin/ml in the plasma. Two days after the start of infusion, all ewes were injected with 100 micrograms cloprostenol and placed with a fertile ram. At slaughter 22 days later, 9 (75%) of the 12 control (saline-infused) ewes were pregnant compared with 1 (11%) of the 9 ewes infused with oxytocin. In the control group, midcycle plasma concentrations of oxytocin were significantly higher in nonpregnant than in pregnant ewes. In Experiment 2, an infertile ram was used throughout to avoid any possible effects of pregnancy and oxytocin infusions were given at different stages of the oestrous cycle. Otherwise the protocol was similar to that in Exp. 1. Oxytocin infusion during luteolysis and the early follicular phase had no effect on the subsequent progesterone secretion pattern, but infusions beginning the day before cloprostenol-induced luteolysis and lasting for 7 or 12 days and infusions beginning on the day of oestrus for 4 days all delayed the subsequent rise in plasma progesterone by approximately 3-4 days. In these animals, the cycle tended to be longer. It was concluded that an appropriate oxytocin secretion pattern may be necessary for the establishment of pregnancy in ewes and that a high circulating oxytocin concentration during the early luteal phase delays the development of the young corpus luteum.     

Rise in plasma beta-endorphin, ACTH, and cortisol in cancer patients undergoing whole body hyperthermia

It has been previously reported that sauna-induced fevers (approximately 39 degrees C) result in rises of beta-endorphins in normal volunteers. This report describes changes in plasma beta-endorphins in cancer patients undergoing whole body hyperthermia (40.5 degrees C to 41.8 degrees C). Results presented show that there is a linear relationship between thermal stress, defined in terms of core temperature and/or duration of hyperthermia, and the quantitative rise in plasma beta-endorphin levels. Data relating to changes in ACTH and cortisol levels are in a single temperature range (41.5 degrees C--41.8 degrees C) are also reported.     

Effect of 3-methylindole on the plasma and lung concentrations of prostaglandins and thromboxane B2 in goats

1. The role of prostanoids in 3-methylindole (3MI)-induced lung disease was investigated. Goats were infused with 3MI in propylene glycol at a dose of 35 mg 3MI/kg body weight. Control goats were infused with propylene glycol alone. 2. Blood was collected at regular intervals starting 24 hr before and ending 72 hr following 3MI infusion. In a second experiment, 3MI-treated goats were killed at 2, 6, 12, 24, 48 and 72 hr post-infusion. The concentrations of PGF2 alpha, PGE, 6-keto PGF1 alpha and TXB2 in plasma and lung of 3MI-infused and control goats were determined by radioimmunoassay. 3. Comparison of individual prostanoid concentrations showed that 3MI-infused and control goats exhibited similar plasma profiles for all four prostanoids measured. 4. In addition, prostanoid concentrations in lungs did not seem to be affected by 3MI infusion. 5. Thus, plasma and lung prostaglandin and TXB2 concentrations do not appear to be altered in 3MI-induced lung disease.     

Febrile and cortisol responses induced in guinea pigs by localized peripheral inflammatory stimulation

(1) In guinea pigs, a high (100 μg/kg) or a low (10 μg/kg) dose of lipopolysaccharide (LPS) was injected into subcutaneously implanted Teflon chambers along with the prior injection of a local anesthetic (ropivacaine) or sterile saline. (2) Intra-chamber injection of the LPS alone induced fever and elevation of circulating cortisol. (3) Fever in response to the low dose of the LPS was attenuated by the pretreatment with the local anesthetic while circulating levels of cortisol were not impaired by this procedure. (4) There was a moderate increase in plasma interleukin-6 (IL-6) in response to both concentrations of locally administered LPS. The LPS did not enter the systemic circulation in measurable amounts. (5) These results favor the possibility of a participation of afferent neural as well as humoral signals (IL-6) in the manifestation of fever in this experimental model.     

Subcutaneous administration of HMG-CoA reductase inhibitors in hyperlipidaemic and normal rats.

Recent reports demonstrate a hypocholesterolaemic effect of daily subcutaneous injections of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors in different rat models of hyperlipidaemia. However, this effect is not seen after oral administration of HMG-CoA reductase inhibitors in rats. We found that oral administration of the HMG-CoA reductase inhibitor Simvastatin also had no effect on plasma cholesterol in severely hyperlipidaemic Nagase analbuminaemic rats (NAR). Simvastatin (an apolar compound dissolved in propylene glycol) was infused continuously for 28 days into the subcutis of control Sprague-Dawley rats (SDR) and NAR using an implanted osmotic pump. All doses which were effective in reducing cholesterol in the NAR (reductions up to approximately 60%), reduced apolipoprotein AI but not apolipoprotein B and caused a severe inflammatory reaction in the dermis. Similar toxicity was observed in the SDR. Subcutaneous administration of the vehicle (propylene glycol) did not cause this reaction and did not affect plasma lipids. Administration of Lovastatin in osmotic pumps resulted in a similar inflammatory reaction. Incorporation of Simvastatin into liposomes did not diminish the toxic effect. On the other hand, infusion of Pravastatin (a polar HMG-CoA reductase inhibitor dissolved in isotonic saline) caused no changes in the dermis and had no effect on plasma lipids in NAR or SDR. Liver microsomes prepared from the Pravastatin-treated rats demonstrated a 3- to 4-fold increase in HMG-CoA reductase activity as compared to untreated rats, confirming uptake of the drug. We conclude that continuous subcutaneous administration of the HMG-CoA reductase inhibitors Simvastatin, Lovastatin and Pravastatin for 28 days may not reduce plasma cholesterol in rats by a mechanism which is related to inhibition of HMG-CoA reductase activity in the liver. The decrease of plasma cholesterol effected by subcutaneous infusion of Simvastatin or Lovastatin in NAR coincides with, and may be related to inflammatory changes caused by administering these compounds into the dermis.     

Infusion of ACTH stimulates expression of adrenal ACTH receptor and steroidogenic acute regulatory protein mRNA in fetal sheep

The late-gestation plasma cortisol surge in the sheep fetus is critical for stimulating organ development and parturition. Increased adrenal responsiveness is one of the key reasons for the surge; however, the underlying mechanisms are not fully understood. Our recent studies suggest that ACTH-mediated increased expression of ACTH receptor (ACTH-R) and steroid acute regulatory protein (StAR) may play a role in enhancing responsiveness. Hence, we examined effects of ACTH infusion in fetal sheep on mRNA expression of these two mediators of adrenal responsiveness and assessed the functional consequences of this treatment in vitro. Fetuses of approximately 118 and 138 days of gestational age (dGA) were infused with ACTH-(1-24) for 24 h. Controls received saline infusion. Arterial blood was sampled throughout the infusion. Adrenals were isolated and analyzed for ACTH-R and StAR mRNA, or cells were cultured for 48 h. Cells were stimulated with ACTH, and medium was collected for cortisol measurement. Fetal plasma ACTH and cortisol concentrations increased over the infusion period in both groups. ACTH-R mRNA levels were significantly higher in ACTH-infused fetuses in both the 118 and 138 dGA groups. StAR mRNA increased significantly in both the 118 and 138 dGA groups. Adrenal cells from ACTH-infused fetuses were significantly more responsive to ACTH stimulation in terms of cortisol secretion than those from saline-infused controls. These findings demonstrate that increases in circulating ACTH levels promote increased expression of ACTH-R and StAR mRNA and are coupled to heightened adrenal responsiveness.     

Local intra-arterial infusion of growth hormone into the mammary glands of sheep and goats: effects on milk yield and composition, plasma hormones and metabolites

Lactating goats and sheep were fitted with catheters in the external pudendal arteries supplying both mammary glands. Saline was infused continuously into one artery whereas the other artery received continuous infusions, over successive 4-day periods, of either saline or growth hormone (GH)-doses increasing twofold between successive periods from 100 to 400 micrograms/day in goats and 400 to 3200 micrograms/day in sheep. Local infusion of GH at up to 1600 micrograms/day in sheep did not affect milk yield or composition nor peripheral plasma concentrations of GH, insulin, glucose, urea and non-esterified fatty acids (NEFA). Infusion of GH at 3200 micrograms/day in sheep increased peripheral plasma concentrations of GH, tended to increase milk yield and peripheral plasma NEFA but there were no changes in peripheral plasma insulin, glucose and urea. It is concluded that GH does not exert direct effects on the mammary glands of sheep and goats in situations where the hormone is administered over short periods.     

Cortisol response of green sturgeon to acid-infusion stress

Cortisol and lactate are classic indicators of stress in fishes and their interactive effects on metabolism during recovery from stress have recently become a subject of more intense study. We examined how stressing green sturgeon through acid infusion affected the cortisol response and lactate metabolism in green sturgeon (Acipenser medirostris). Both lactic acid (0.3 M) and HCl (0.3 N) infusion (infusion volumes 1.5 ml kg(-1)) elicited an immediate cortisol response (21.61+/-4.61 ng ml(-1) and 17.50+/-3.00 ng ml(-1), respectively). Lactic acid prolonged the cortisol response compared to HCl (90 min vs. 25 min). Neutralized lactate (0.23 M; with 1 N NaOH; final pH 7.8) and NaCl (0.9%) infusion (infusion volumes 1.5 ml kg(-1)) did not affect plasma cortisol. Sturgeon infused with lactic acid showed a faster rate of lactate disappearance from plasma than those with neutralized lactic acid. We relate these findings to lactate metabolism following exercise, acid-infusion and air immersion stress in fishes.     

Characterization of the febrile response induced by fibroblast-stimulating lipopeptide-1 in guinea pigs

Recently, it has been shown that the Toll-like receptors-2 and -6 agonist fibroblast-stimulating lipopeptide-1 (FSL-1) have the capacity to induce fever and sickness behavior in rats. Since the mechanisms of the fever-inducing effects of FSL-1 are still unknown, we tested the pyrogenic properties of FSL-1 in guinea pigs and assessed a role for TNF-alpha and prostaglandins in the manifestation of the febrile response to this substance. Intra-arterial and intraperitoneal injections of FSL-1 caused dose-dependent fevers that coincided with elevated plasma levels of TNF and IL-6, the intraperitoneal route of administration being more effective than the intra-arterial route. Intra-arterial or intraperitoneal injection of a soluble form of the TNF type 1 receptor, referred to as TNF binding protein (TNFbp), together with FSL-1, completely neutralized FSL-1-induced circulating TNF and reduced fever and circulating IL-6. Intra-arterial or intraperitoneal injection of the nonselective cyclooxygenase (COX)-inhibitor diclofenac depressed fever and FSL-1-induced elevations of circulating PGE2. Circulating TNF and IL-6, however, remained unimpaired by treatment with diclofenac. In conclusion, FSL-1-induced fever in guinea pigs depends, in shape and duration, on the route of administration and is, to a high degree, mediated by pyrogenic cytokines and COX products.