Jeffrey L. Selph

Some quantitative temporal characteristics of carrageenin-induced pleurisy in the rat.

Summary The intrapleural injection of 500 μg of carrageenin produces an acute inflammatory reaction in the rat in which it is possible to recover quantitatively the inflammatory cells that are mobilized and the exudate. The time courses of exudate formation and neutrophil mobilization strongly suggest that these phenomena are related. Both curves are sigmoid, and increase sharply between the first and third hours. Aspirin at 90 mg/kg, p.o., produced 83% inhibition of exudate formation at three hours but by the seventh hour the inhibition was only 31%. Neutrophil mobilization was only slightly inhibited by aspirin at three hours. However, the neutrophil mobilization which occurred between the third and seventh hours was completely blocked by aspirin. Monocyte mobilization started two hours after injection of the carrageenin and continued at a fixed rate through the seventh hour. There was no mobilization of monocytes in the aspirin-treated animals for the first five hours. Thereafter the rate of mobilization of these cells was similar in control and aspirin-treated animals. Three lines of evidence suggest that the single period of edema formation produced by the intrapleural injection of carrageenin in the first 5 hr is analogous to the second phase of the biphasic rat hindlimb reaction to this irritant. The major portion of these edemas is formed 1–3 hr after injection of carrageenin. Temporally associated with the development of both edemas is the mobilization of large numbers of neutrophils and the appearance of these cells free in the inflamed tissue. Finally, both edemas are strongly inhibited by relatively low doses of aspirin and both are insensitive to the antihistaminic triprolidine and the antiserotonin agent cyproheptadine. The authors wish to thank Dr. Robert A. Maxwell, Dr. Donald H. Namm, and Dr. Helen L. White for their advice and constructive criticism in the preparation of this manuscript.

Quantitative comparison of the analgesic and anti-inflammatory activities of aspirin, phenacetin and acetaminophen in rodents

The mild analgesic activities of aspirin, phenacetin and acetaminophen have been compared in the trypsin, kaolin and carrageenan hyperalgesic assays as well as in the acetic acid writhing test. The trypsin and kaolin hyperalgesic assays were designed to be unaffected by drugs with anti-inflammatory activity. Aspirin and acetaminophen were inactive in these two tests at dose levels devoid of side effects. Phenacetin was active in the trypsin and kaolin assays with oral ED50s of 114 +/- 36.2 and 107 +/- 11.5 mg/kg, respectively. Non-steroidal anti-inflammatory drugs as well as phenacetin and acetaminophen were active in the acetic acid writhing and carrageenan hyperalgesic assays. This led to evaluation of phenacetin and acetaminophen as anti-inflammatory agents. Both of these latter drugs were active in the carrageenan pleurisy and adjuvant arthritis models of inflammation. In all studies phenacetin was equipotent to or more potent than acetaminophen. The data suggest that the analgesia produced by aspirin and acetaminophen results from their anti-inflammatory activity whereas the analgesia produced by phenacetin has two components, one dependent on and one independent of anti-inflammatory activity.

Potentiation of the Anti-Inflammatory and Analgesic Activity of Aspirin by Caffeine in the Rat

Summary Caffeine has been found to potentiate the acute anti-inflammatory activity of aspirin, indomethacin, and phenylbuta-zone, but not the activity of sodium salicy-late or hydrocortisone, in the carrageenan pleurisy or hindlimb models of inflammation in the rat. The mobilization of inflammatory cells was not affected by aspirin in the presence or absence of caffeine. The mild analgesia produced by aspirin was confined to a hyperalgesic test in which this drug was able to reduce inflammation and concomitant hyperalgesia and thereby produce an “apparent” analgesic effect. This “apparent” analgesia produced by aspirin was potentiated by caffeine. The mechanism responsible for the potentiated anti-inflammatory and mild analgesic activity of aspirin remains unknown since caffeine did not alter the plasma salicylate levels or prostaglandin synthetase inhibition produced by aspirin. The authors wish to thank Dr. R. A. Maxwell and Dr. D. H. Namm for their advice and constructive criticism in the preparation of this manuscript.

New analgesic assay utilizing trypsin-induced hyperalgesia in the hind limb of the rat

Subplantar injection of 250 micrograms of trypsin in the rat resulted in a biphasic increase in pain sensitivity (hyperalgesia) with peaks at 10 and 150 min separated by a period of decreased sensitivity to pain (hypoalgesia). Hyperalgesia was assessed by a decrease in response latency to a 3.0-kg force applied to the injected hind limb. Response latencies at 150 min were increased in a dose-dependent manner by pretreatment at 90 min with acetaminophen; phenacetin; the arachidonate cyclooxygenase inhibitors aspirin, indomethacin, and ibuprofen; and the opiate analgesics codeine and morphine. ED50s of 17, 13, 10, 0.48, 1.6, 3.9 and 1.2 mg/kg p.o. were obtained for these drugs, respectively. The hyperalgesia present at 150 min was not affected by pretreatment with antiinflammatory steroids, an antihistaminic, an antiserotonin agent, and an anticholinergic. We recommend measurement of drug-induced increase in response latencies produced 150 min after injection of 250 micrograms of trypsin as the basis for a new sensitive and selective analgesic assay. ED50s obtained in this assay correlate well with doses that are used clinically to produce analgesia. Development of the hypoalgesic component was selectively inhibited by pretreatment with an antiserotonin agent. Additional drug studies indicated that the algesic response to the subplantar injection of trypsin is the resultant of independent, temporally overlapping hyperalgesic and hypoalgesic components.

Arachidonate metabolic pathways in cells harvested from rat pleural cavity at various times after carrageenan administration

Cells were harvested from rat pleural cavity before and during the inflammatory response stimulated by carrageenan injection. The conversion of [14C]arachidonate by intact cells into products of the cyclooxygenase and 5-lipoxygenase pathways was studied in the absence and presence of ionophore. Incorporation of arachidonate into phosphatidic acid was also followed. In the absence of ionophore, the principal arachidonate metabolites of resident macrophages were the cyclooxygenase products, prostacyclin and thromboxane, while mobilized monocytes produced thromboxane as a major product, but very little prostacyclin. Arachidonate-metabolizing enzymes in mobilized monocytes were, in general, less active than those of resident macophages. Cells harvested at times when mobilized neutrophils were the predominant cell type were capable of converting arachidonate into thromboxane and prostaglandins, but not prostacyclin. These cells exhibited the most active turnover of arachidonate into phosphatidic acid, and the extent of this turnover appeared to be temporally related to the presence of edema in the pleural cavity at the time of cell harvest. Enzymatic formation of 5-lipoxygenase products was dependent on calcium and was markedly stimulated by ionophore A23187 in both resident and mobilized pleural cells. Among several non-steroidal drugs tested, cyclooxygenase inhibitors were the most effective in preventing the inflammatory response in the carrageenan model of inflammation.

Pharmacological characterization of the algesic response to the subplantar injection of serotonin in the rat

Subplantar injection of 0.10 micrograms of serotonin in the rat resulted in a brief period (0-20 min) of increased pain sensitivity to an applied force (hyperalgesia) which preceded a longer period (40-120 min) of decreased pain sensitivity (hypoalgesia). The magnitude of each of these changes and the duration of the hypoalgesia were dose-dependent. The development of hyperalgesia was selectively and dose dependently reduced by inhibitors of arachidonate cyclooxygenase. The hypoalgesia was selectively and dose dependently reduced by the serotonin antagonist methysergide. Selective inhibition of the hyperalgesia by aspirin and of the hypoalgesia by methysergide revealed that components of both hyperalgesia and hypoalgesia were present in the 10-120 min interval. These findings, the level of serotonin reported to be released in rat dermal tissue, and selective drug inhibition studies suggest that some irritant-induced changes in algesia measured in the rat hindlimb result from release of dermal stores of serotonin. Selective inhibition of the hypoalgesic component of the hindlimb irritant trypsin by the antiserotonin agent methysergide supports this hypothesis. The principal conclusion derived from these studies is that the algesic response to the subplantar injection of a single agent can be the resultant of independent, temporally overlapping hyperalgesic and hypoalgesic components each of different intensity and pharmacological sensitivity.

Development of Carrageenan Pleurisy in the Rat: Effects of Colchicine on Inhibition of Cell Mobilization

Abstract Colchicine produced three effects which modified the acute inflammatory response to carrageenan in the rat pleural cavity: (i) inhibition of neutrophil mobilization and concomitant exudate formation (3 hr); (ii) inhibition of monocyte mobilization (21 hr); and (iii) augmented exudate formation (3 and 21 hr). The 1st effect was related to the intraperitoneal dose of colchicine and occurred only at leukopenic dose levels. This effect could not be produced by intrapleural injection of nonleukopenic doses of colchicine. The second effect, on the other hand, was produced by intraperitoneal leukopenic doses and to a lesser extent by intrapleural administration of nonleukopenic doses of colchicine. Importantly, the normal biphasic exudative response to carrageenan developed fully in the absence of monocytes. The third effect, a dose-dependent augmentation of both exudative phases of carrageenan pleurisy, was produced by low, nonleukopenic, intrapleural doses of colchicine. The augmented exudate was sensitive to prostaglandin synthetase inhibitors but not to anti-inflammatory steroids. Neither neutrophils nor monocytes were responsible for the augmented exudate. Colchicine, injected into the rat hindlimb or pleural and peritoneal cavities did not elicit the mobilization of neutrophils or a pleural effusion. In addition, colchicine did not affect the magnitude, temporal development, or decay of the potent edematogenic action of serotonin in the rat hindlimb. Thus irritancy was not responsible for any of the effects of colchicine.

Azathioprine Treatment of Adjuvant Arthritis

Administration of azathioprine in the diet to Lewis rats reduced the lethality of this drug relative to various regimens by gavage. This mode of administration made it possible to administer effective doses with reduced toxicity. Azathioprine blocked the early development of adjuvant arthritis and decreased the joint scores of animals with established adjuvant disease. A combination of azathioprine and prednisolone produced an additive reduction of both developing and established joint scores. A 10 day pulse regimen of prednisolone resulted in a strong, rapid decrease in established joint scores. This decrease was sustained by continuous administration of azathioprine. In all cases removing azathioprine and/or prednisolone from the diet of animals with established adjuvant arthritis resulted in recurrence of disease. The results of these studies support the validity of the adjuvant model for prediction of anti-arthritic activity.