The pharmacology of fever.

The ability to minimise, if not prevent, large variations in deep body temperature that would otherwise result from some environmental conditions is a homeostatic function of unquestioned benefit that is demonstrated only by the more highly evolved animals. Nevertheless, body temperature is raised above normal values in many pathological conditions. This increase in temperature or fever is an active and co-ordinated response, which indicates the involvement of the CNS. Central injection and lesion studies have shown that the brain, in particular the PO/AH, is the site of action of fever-inducing agents, termed pyrogens. Electrophysiological data show that pyrogens modify the activity of central thermosensitive neurones as if to increase heat gain and decrease heat loss. The common response of fever to pyrogens of diverse origins is attributable to fever being mediated by an endogenous pyrogen released by phagocytic cells in the host. The mechanism by which central neuronal function is disturbed by pyrogens present in the periphery is not known. Tracer studies have yet to demonstrate the passage of a pyrogen across the blood-brain barrier. The possible involvement of several putative neurotransmitters and modulators in fever has been reviewed here, but most compounds have not been studied sufficiently to allow firm conclusions to be drawn. Much of the data is limited to the effects of the putative mediators on normal thermoregulation but, even when the effect is hyperthermia, such observations do not necessarily indicate a role for the endogenous material in fever. Dose-response curves for agonists and the effects of antagonists are often undetermined. This shortfall in data is due to some extent to the nature of fever; a central response in vivo over several hours. Although fever may enhance other host reactions to combat infection and inflammation, neither this benefit nor the undesirability of antipyretic therapy has been demonstrated unequivocally in either homeothermic laboratory animals or humans. Consequently, antipyretic drugs continue to be used clinically to alleviate the fever, malaise and/or pain commonly associated with disease. The drugs in common usage are the nonsteroidal antipyretic analgesics, many of which also have an anti-inflammatory effect. The primary mode of action of these drugs as antipyretics appears at present to be the inhibition of cyclo-oxygenase and a consequent reduction of prostanoid material in pyrogen-sensitive areas of the brain.(ABSTRACT TRUNCATED AT 400 WORDS)

Evidence against adenosine 3',5'-monophosphate as a mediator of fever in the brain.

The aim of this study was to test the hypothesis that increased concentrations of adenosine 3',5'-monophosphate (cyclic AMP) in the pyrogen-sensitive preoptic/anterior hypothalamic region (PO/AH) mediate fever. Micro-injection of N6-2'-O-dibutyryl adenosine 3',5'-monophosphate (db cyclic AMP) into the preoptic/anterior hypothalamic region in rats produced a dose-dependent fall in body temperature which is inconsistent with the proposal that the nucleotide mediates fever. Hyperthermia was observed in some rats in response to large doses of db cyclic AMP, but this response was associated with convulsions. Endogenous concentrations of cyclic AMP in the preoptic/anterior hypothalamic region, as well as in the cerebral cortex, liver, spleen, thymus, white fat and plasma were unaffected by the febrile response to the subcutaneous injection of yeast in rats. A rise in levels of cyclic AMP was observed in the skeletal muscle of rats treated with yeast. The data presented do not indicate that cyclic AMP is involved in the neuronal events mediating fever in the rat.

Rational drug design approach for overcoming drug resistance: application to pyrimethamine resistance in malaria.

Pyrimethamine acts by selectively inhibiting malarial dihydrofolate reductase-thymidylate synthase (DHFR-TS). Resistance in the most important human parasite, Plasmodium falciparum, initially results from an S108N mutation in the DHFR domain, with additional mutation (most commonly C59R or N51I or both) imparting much greater resistance. From a homology model of the 3-D structure of DHFR-TS, rational drug design techniques have been used to design and subsequently synthesize inhibitors able to overcome malarial pyrimethamine resistance. Compared to pyrimethamine (Ki 1.5 nM) with purified recombinant DHFR fromP. falciparum, the Ki value of the m-methoxy analogue of pyrimethamine was 1.07 nM, but against the DHFR bearing the double mutation (C59R + S108N), the Ki values for pyrimethamine and the m-methoxy analogue were 71.7 and 14.0 nM, respectively. The m-chloro analogue of pyrimethamine was a stronger inhibitor of both wild-type DHFR (with Ki 0.30 nM) and the doubly mutant (C59R +S108N) purified enzyme (with Ki 2.40 nM). Growth of parasite cultures of P. falciparum in vitro was also strongly inhibited by these compounds with 50% inhibition of growth occurring at 3.7 microM for the m-methoxy and 0.6 microM for the m-chloro compounds with the K1 parasite line bearing the double mutation (S108N + C59R), compared to 10.2 microM for pyrimethamine. These inhibitors were also found in preliminary studies to retain antimalarial activity in vivo in P. berghei-infected mice.

Evidence that cyclic nucleotides are not mediators of fever in rabbits.

The N6-2'-O-dibutyryl derivative of adenosine 3',5'-monophosphate (db cyclic AMP) and related compounds have been micro-injected into the preoptic/anterior hypothalamic nuclei (PO/AH) of the unanaesthetized, restrained rabbit and the effects on deep body temperature observed. Db cyclic AMP (100-400 micrograms) produced hypothermia of rapid onset in rabbits at an ambient temperature of 20-23 degrees C. Hypothermia was also produced by N2-2'-O-dibutyryl guanosine 3',5'-monophosphate (db cyclic GMP), but not by saline, sodium n-butyrate, adenosine 3',5'-monophosphate (cyclic AMP), guanosine 3',5'-monophosphate, adenosine 5'-mono-, di- or triphosphate. The initial hypothermic response to db cyclic AMP and db cyclic GMP was followed by a sustained rise in temperature. However, all compounds injected into the PO/AH produced a similar hyperthermia which was attenuated by paracetamol. Development of this tissue-damage fever abolished the hypothermic response to db cyclic AMP in some rabbits. The effects of db cyclic AMP on body temperature and behaviour were not reproduced by the adenylate cyclase activators, cholera toxin (0.125-5 micrograms) and guanyl imidodiphosphate (5-400 micrograms). It is concluded that hypothermia is the principal effect of db cyclic AMP on body temperature when injected into the PO/AH in rabbits. These data do not support the proposal that endogenous cyclic AMP in the rabbit brain mediates pyrexia.

Study on the possible entry of bacterial endotoxin and prostaglandin E2 into the central nervous system from the blood.

1 A study has been made of the possible entry of 51Cr-bacterial endotoxin and [5,6,8,11,12,14,15(n)-3H]-prostaglandin E2 ([3H5-PGE2) into the CNS of the anaesthetized cat. 2 No radioactivity was detected in perfusates of the preoptic-anterior hypothalamus or in the cerebrospinal fluid (c.s.f.) in vivo, or in brain tissue post mortem following intracarotid infusion of 51Cr-bacterial endotoxin. 3 Intracarotid administration of [3H]-PGE2 resulted in the entry of radioactivity into the CNS of endotoxin pretreated cats. Chromatographic analysis indicated the radioactivity in c.s.f. to be associated with PGE2 and a metabolite similar to 13, 14-dihydro-15-keto PGE2. 4 Intracarotid administration of 13, 14-dihydro-15-keto [5,6,8,11,12,14(n)-3H]-PGE2 resulted in the presence of the compound in the CNS of the anaesthetized cat after pretreatment with bacterial endotoxin. 5 It is concluded that PGE2 and possibly 13,14-dihydro-15-keto PGE2 but not bacterial endotoxin may enter the CNS from the cerebral circulation to elicit the febrile response to bacterial endotoxin in cats.

Dissimilar effects on body temperature in the cat produced by guanosine 3',5'-monophosphate, acetylcholine and bacterial endotoxin.

1 Guanosine 3',5'-monophosphate (cyclic GMP) and N2-2'-O-dibutyryl guanosine 3',5'-monophosphate (db cyclic GMP) have been injected into the third cerebral ventricle (i.c.v.) of the unanaesthetized cat and the effects of rectal temperature and on behavioural and autonomic activities observed and compared with those of acetylcholine and physostigmine. 2 Acetylcholine (100 nmol) and physostigmine (100 nmol) injected together i.c.v. produced a rise in body temperature in cats at an environmental temperature of 20-24 degrees C, which was abolished by pretreatment i.c.v. with atropine (200 nmol). 3 Cyclic GMP and db cyclic GMP (10--1250 nmol) had no effect on body temperature in cats at an environmental temperature of 20--24 degrees C but produced hypothermia (1250 nmol) in cats at an environmental temperature of 9--11 degrees C. 4 The O-somatic antigen of Shigella dysenteriae (20 microgram/kg i.v.) produced fever in cats which was not potentiated by caffeine (25 mg/kg i.p.). Levels of endogenous cyclic GMP in c.s.f. taken from the cisterna magna during fever induced by bacterial endotoxin in the presence or absence of paracetamol (50 mg/kg i.p.) and/or caffeine were similar to values for afebrile cats. 5 It is concluded that exogenous cyclic GMP and db cyclic GMP can inhibit central events mediating autonomic and behavioural thermoregulation stimulated in cats by exposure to cold environments.

Thermoregulatory effects of N6-2'-Q-dibutyryl adenosine 3',5'-monophosphate in the restrained mouse.

1 The N6-2-O-dibutyryl derivative of adenosine 3',5'-monophosphate (db cyclic AMP) has been micro-injected into the third cerebral ventricle of the unanaesthetized, restrained mouse and the effects on body temperature and thermoregulatory activities observed. 2 Db cyclic AMP (4, 16 and 32 micrograms) injected intracerebroventricularly produced hypothermia when compared with temperature responses to sodium n-butyrate (6.8 micrograms). 3 Hypothermia induced by db cyclic AMP in mice was associated with a fall in oxygen consumption together with behavioural and autonomic heat loss activities but not cutaneous vasodilatation. The effects on rectal temperature and oxygen consumption were dose-dependent. 4 The falls in rectal temperature and oxygen consumption induced by db cyclic AMP (4 micrograms) were decreased by elevation of the environmental temperature from 22 to 32 degrees C and abolished at 36 degrees C. 5 It is concluded db cyclic AMP may inhibit central events mediating the rise in metabolic heat production in mice upon exposure to cold environments.

Effects of malaria on O2 consumption and brown adipose tissue activity in mice.

Increased energy expenditure often occurs during illness or after injection of endotoxin and can contribute to the generation of fever. In laboratory rats and mice the thermogenic response has been attributed to the sympathetic activation of brown adipose tissue (BAT), although mice often fail to show pyrexia. In this study the effects of malaria on O2 consumption and BAT were studied in mice inoculated with Plasmodium berghei. Parasitemia was maximal (greater than 50% of erythrocytes showing positive Leishman staining) 72 h after inoculation. Up to this time body weight and food intake were similar to values for control mice, although colonic temperatures were slightly depressed in infected mice. Thereafter, infected mice showed marked hypophagia, loss of body weight, and severe hypothermia; colonic temperature was less than 31 degrees C at 96 h when the experiment was terminated. Resting O2 consumption (VO2) measured at 24 degrees C was slightly elevated in infected mice 12 h after inoculation and reached a peak value (31% above controls) at 48 h. VO2 returned to the same value as controls at 96 h. In vitro thermogenic activity of BAT (assessed from binding of guanosine diphosphate to isolated mitochondria) was not significantly altered in infected mice. These data demonstrate a marked thermogenic response to malarial infection, but this is not accompanied by fever in mice and is dissociated from stimulation of BAT activity.

Nitroglycerine reduces neutrophil activation and acute damage in latissimus dorsi muscle grafts.

BACKGROUND: Damage to the latissimus dorsi muscle (LDM) may jeopardize a successful outcome to dynamic cardiomyoplasty. We and others have demonstrated muscle damage in LDM in various species including humans. Ischemia is now recognized to be an important contributory factor. We postulated that glyceryl trinitrate, a nitric oxide donor, might protect against ischemic endothelial dysfunction and so reduce resultant muscle damage. METHODS: In 20 adult rats the left LDM was mobilized on its thoracodorsal neurovascular pedicle and maintained as an orthotopic graft. Half of the animals received glycerol trinitrate intraoperatively and postoperatively for 24 hours. The other half served as untreated controls. Each group was further subdivided into two groups (n = 5 in each): animals in which the LDM was excised after 4 hours for myeloperoxidase studies, and animals in which the LDM was excised at 24 hours for analysis of muscle damage by histology and enzyme macrohistochemistry. Blood samples were taken at 24 hours for assay of plasma nitrite and nitrate as nitric oxide metabolites. RESULTS: Glycerol trinitrate-treated animals had higher plasma nitric oxide metabolite levels after 24 hours (after nitrate reductase treatment, total nitrite, 78.3+/-11.8 nmol/mL, mean +/- SEM) than controls (42.1+/-3.7 nmol/ mL, p = 0.008). The proportion of viable LDM in glycerol trinitrate-treated animals was greater than in untreated animals, mainly in the middle and distal regions of the graft (middle region, 96.3%+/-0.5% versus 75.7%+/-4.1%, p<0.001; distal region, 94.4%+/-0.8% versus 40.9%+/-3.1%, p<0.001). Macrohistochemical findings correlated well with the histologic findings. Myeloperoxidase activity (U/g) was markedly lower in glycerol trinitrate-treated LDMs, mainly in the distal part of the graft (glycerol trinitrate versus control, 20.5+/-2.1 versus 40.9+/-3.1 U/g, p<0.001). CONCLUSIONS: Glycerol trinitrate significantly reduced acute damage to the distal two-thirds of the mobilized LDM, possibly by modifying leukocyte activation and endothelial dysfunction associated with ischemic injury.

Effects of methylxanthine drugs on pyrogen-induced hyperthermia.

The hyperthermic response to pyrogen was not potentiated by caffeine or theophylline administered i.p. into cats or rabbit. Injection of these drugs into the anterior hypothalamus or into the third cerebral ventricle in cats was also without effect on pyrexia. These results support the hypothesis that cyclic AMP in the anterior hypothalamus does not mediate pyrogen-induced hyperthermia in cats.

Effects of diflunisal on fever in the rabbit and the rat.

The antipyretic efficacy of diflunisal was assessed in rats made febrile by yeast and in rabbits made febrile by bacterial endotoxin. Diflunisal was a more potent antipyretic than aspirin in rats, reducing a maximum fever in doses not producing overt toxic effects. In contrast, submaximal fever in rabbits was not reduced by diflunisal. Fatal hyperthermia of rapid onset was observed in rats and rabbits receiving high doses of diflunisal after administration of microbial pyrogen but not in control animals. These data indicate the toxicity of diflunisal may be potentiated by the presence of pyrogens. It is concluded that the apparent antipyretic efficacy of a drug can depend on the species-pyrogen combination used to screen for antipyresis.

Evidence against putrescine and polyamines as endogenous mediators of fever.

Endogenous concentrations of putrescine, spermidine, spermine and related biosynthetic enzymes were not affected by the administration of bacterial endotoxin and the subsequent development of fever in rabbits. In addition, the febrile response to endotoxin was unaffected either by the ornithine decarboxylase inhibitor, DL-alpha-difluoromethylornithine or by putrescine. These data indicate polyamines are not involved in the development of fever.

Ergometrine and 5-hydroxytryptamine binding sites in rat brain and myometrium.

Functional studies suggest that ergometrine is a partial agonist involving 5-hydroxytryptamine (5-HT) receptors in rat uterus. Ergometrine displaced [3H]5-HT from specific binding sites in rat brain, but did not displace [3H]5-HT at functionally important concentrations in rat myometrium. These binding studies indicate that the agonist and antagonist actions of ergometrine in rat uterus arise from its initial interaction with binding sites other than those for 5-HT.

An exploration of the structure-activity relationships of 4-aminoquinolines: novel antimalarials with activity in-vivo.

The structure-activity relationships of bisquinolines, a potentially important group of novel antimalarial drugs, were studied. The high-temperature (180-250 degrees C) synthesis of 4-aminoquinolines, including bisquinolines, by nucleophilic displacement was both fast and efficient Several bisquinolines including (+/-)-trans-N1,N2-bis(7-trifluoroquinolin-4-yl)cyclohexane-1, 2-diamine and 1R,2R-(-)-, 1S,2S-(+)-, (+/-)-trans- and cis-N1, N2-bis(7-chloroquinolin-4-yl)cyclohexane-1,2-diamine exhibited potent activity against Plasmodium berghei in mice; (+/-)-trans-N1,N2-bis(7-chloroquinolin-4-yl)cyclohexane-1, 2-diamine was orally active. Our results indicate that these compounds conform to a putative receptor for quinoline antimalarials. In addition, a 7-haloquinoline linked by a heterocyclic bridge, at the 4-position, to another heterocycle (such as an acridine at the 9-position) maximally occupies the active site of our postulated target.

Novel aryl-bis-quinolines with antimalarial activity in-vivo.

Three rationally designed isomeric aryl-bridged bis-quinolines, N1,Nx-bis(7-chloroquinolin-4-yl)phenylene-1,x-diamines, where x=2, 3 or 4, i.e. o-, m- and p-substituted analogues respectively, were synthesized and evaluated against Plasmodium berghei in-vivo. The compound with x=2 had an ID50 of 30 mg kg(-1), whereas the p-substituted analogue (x=4) was not statistically schizonticidal at either of the two dose levels tested in olive oil-dimethylsulphoxide (5 and 25 mg kg(-1), ID50=60 mg kg(-1) approx.). When the delivery vehicle was changed to saline-DMSO, antimalarial potency increased for the p-substituted compound (ID50 17 mg kg(-1)). In contrast, the m-substituted analogue had marked antimalarial activity (ID50 1.2 mg kg(-1)), which compares favourably with that of chloroquine diphosphate (ID50 = 4.3 mg kg(-1)). The data presented show that the aminomethylene side chain in amodiaquine can be successfully replaced by a 7-halo-4-aminoquinoline, establishing that carbon bridges containing less than four contiguous carbon atoms can be present within highly active aryl-substituted 4-aminoquinoline antimalarials. These results confirm that the presence of an OH group in the aryl bridge is not necessary for antimalarial activity and substantiate the view that, despite the appearance of resistant strains, new and existing aminoquinolines still have an important role in treating malaria.

Cyclic adenosine 3', 5'-monophosphate in cerebrospinal fluid during thermoregulation and fever.

1. Samples of cerebrospinal fluid (c.s.f.) have been taken from the cisterna magna of unanaesthetized cats, whilst rectal temperature was recorded, during exposure of the animals to various ambient temperatures and during fever induced by pyrogen. The concentration of adenosine 3', 5'-monophosphate (cyclic AMP) in samples of c.s.f. has been assayed. 2. Cats exposed to low ambient temperatures (-2 to +2 degrees C) for 3 h maintained body temperature by both behavioural and autonomic heat gain activity. Exposure of cats to high ambient temperatures (44 - 45 degrees C) for 3.5 h caused a rise in body temperatures of about 2.5 degrees C, despite behavioural and autonomic heat loss activity. Neither cold nor heat stress had a significant effect on c.s.f. cyclic AMP. 3. Fever induced by intravenous Shigella dysenteriae (2 and 20 mug/kg) was associated with a dose-related increase in the concentration of cyclic AMP in c.s.f. Paracetamol (75 mg/kg) injected I.P. before the onset of fever, suppressed the increase in both temperature and c.s.f. cyclic AMP in response to pyrogen. Paracetamol (50 and 100 mg/kg), injected after the onset of fever, caused a fall in temperature, which was not associated with a decrease in the concentration of cyclic AMP in c.s.f. 4. Fever induced in cats by intravenous Shigella dysenteriae (20 mug/kg) was associated with an increase in the concentration of cyclic AMP in plasma as well as in c.s.f. 5. The sodium salt of cyclic AMP (0.1-10 mg/kg) injected I.V. into unanaesthetized cats caused a dose-related hypothermia, which was associated with autonomic heat loss activity and a dose-related increase in the concentration of cyclic AMP in cisternal c.s.f., which was not mimicked by adenosine. 6. It is concluded that the raised concentrations of cyclic AMP in c.s.f., in response to pyrogen I.V., do not mediate fever in the cat and that the concentration of cyclic AMP in cisternal c.s.f. may be affected by changes in the plasma concentration of the nucleotide.

Nitric oxide and reactive nitrogen intermediates during lethal and nonlethal strains of murine malaria.

The virulence of Plasmodia depends partly on the strain of parasite and partly on the host. In this study, Plasmodium berghei N/13/1A/4/203 caused the death of mice, whereas Plasmodium chabaudi chabaudi AS was not lethal. Current opinion is that nitric oxide (NO) and other reactive nitrogen intermediates (RNI) are produced in several host organs during malaria to resist infection or produce tissue damage. NO and RNI production in blood or plasma, brain, liver and spleen in MF1 mice was investigated during P. berghei and P. c. chabaudi infection, in order to help determine whether changes in NO production are beneficial or detrimental to the host in vivo. NO production was measured both directly and indirectly as nitrites and nitrates, to represent RNI. No changes in blood NO were detected in P. berghei infected mice, but increases were observed in brain, liver and spleen. In P. c. chabaudi infected mice, rises in NO concentration were observed in blood and spleen, whereas a decline in liver NO was seen, but there were no changes in brain. Liver contained the highest concentration of RNI, but increasing concentrations were seen in both plasma and spleen in both P. berghei and P. c. chabaudi infected mice. These results show that NO and RNI production alters during murine malaria. The changes depend upon the tissue, the day of infection, the degree of parasitaemia, the strain of Plasmodia and the method of measuring NO biosynthesis. Lethal P. berghei induced NO production in the mid and late stages of infection in mice when parasitaemia was high, whereas in nonlethal P. c. chabaudi infection, NO production was increased in the early and late stages when parasitaemia was low. These data are consistent with a role for NO in the protection of the MF1 mouse against Plasmodia. Failure to clear the parasite is associated with evidence of increased NO production in brain and liver, which may contribute to the pathology of malaria, but this hypothesis requires confirmation from other experimental approaches.