Enhancement of neutrophil infiltration in histidine decarboxylase-deficient mice

The roles of histamine in the anaphylactic increase in vascular permeability and leucocyte infiltration were analysed in an air pouch-type allergic inflammation model in histidine decarboxylase-deficient (HDC^−/−) mice and wild-type mice. In the immunized wild-type mice, histamine content in the pouch fluid and vascular permeability in the anaphylaxis phase were increased by injection of the antigen solution into the air pouch. However, in the immunized HDC^−/− mice, the antigen challenge did not increase histamine content in the pouch fluid and vascular permeability in the anaphylaxis phase. Number of leucocytes (more than 83% are neutrophils) in the pouch fluid 4–24 hr after the antigen challenge in the HDC^−/− mice was significantly higher than that in the wild-type mice. Simultaneous injection of histamine with the antigen solution into the air pouch of the immunized HDC^−/− mice reduced the antigen-induced leucocyte infiltration at 4 hr. Simultaneous injection of the H2 antagonist cimetidine but not the H1 antagonist pyrilamine with the antigen solution into the air pouch of the immunized wild-type mice further increased leucocyte infiltration at 4 hr. The levels of macrophage inflammatory protein-2 at 2 hr and of tumour necrosis factor-α at 4 hr in the pouch fluid of the HDC^−/− mice were significantly higher than those of the wild-type mice. These findings indicate that histamine plays significant roles not only in the anaphylactic increase in vascular permeability via H1 receptors but also in the negative regulation of neutrophil infiltration via H2 receptors in allergic inflammation.     

Characterization of methylated bovine serum albumin-induced allergic inflammation in rats.

An air pouch type allergic inflammation in rats was induced using an insoluble cationic protein, methylated bovine serum albumin (MeBSA), as an antigen. Changes in vascular permeability, local tissue edema, histamine contents in the pouch fluid, and number of infiltrated leukocytes and chemotactic activity in the pouch fluid were analyzed during an 8-hour period after injecting the antigen solution into the air pouch of the immunized and nonimmunized rats. Vascular permeability during the first 30-min interval in the immunized rats was higher than that in the nonimmunized rats, reflecting a higher histamine level in the pouch fluid. However, both the increase in vascular permeability and histamine level in the immunized rats in this period were much lower than those induced by a soluble, noncationic antigen, azobenzenearsonate-conjugated acetyl bovine serum albumin. In the MeBSA-induced allergic inflammation model, a second peak of vascular permeability was induced at 2 h, and local tissue edema formation became apparent at 2 h, reaching a plateau at 4 h. A prominent increase in leukocyte infiltration, especially neutrophils, into the pouch fluid was induced at 4 h in accordance with an increase in chemotactic activity in the pouch fluid. These observations indicate that the acute phase of MeBSA-induced allergic inflammation is characterized by a weak anaphylactic response and a prominent neutrophil infiltration.     

Inhibitory effect of indomethacin farnesil,a novel antiinflammatory prodrug,on carrgeenin-induced inflammation in rats

Antiinflammatory effects of indomethacin farnesil (IMF), a novel prodrug of indomethacin, was examined after both oral and local administration. In the air pouch carrageenin-induced inflammation, an oral dose of IMF exerted dose dependent inhibitory effects on the accumulation of inflammatory exudate fluid and the migration of leukocytes into the exudate. Both the increased vascular permeability and the prostaglandin E₂ levels in the exudate fluid were reduced by IMF. Significant levels of free indomethacin were detected in the pouch fluid. In spite of the inability of IMF to inhibit prostaglandin synthesis in a cell free cyclooxygenase system, IMF injected locally inhibited carrageenin paw edema, and the inhibitory effect was comparable to that of indomethacin itself. When injected locally into the paw together with carrageenin,¹⁴C-IMF was effectively converted to its active metabolite, indomethacin. The indomethacin concentration in the paw tissue was comparable to that of indomethacin injected paws with the same molar dose of free indomethacin.     

Increase in vascular permeability induced by leukotriene b4 and the role of polymorphonuclear leukocytes

Leukotriene B₄ (LTB₄), a metabolite of arachidonic acid, is known to be a potent chemotactic and chemokinetic substance. We have used the hamster cheek pouch microcirculation model to study the effect of LTB₄ on vascular permeability and the involvement of neutrophil granulocytes in this response. Intravascular fluorescein-labeled dextran (mol wt 150,000) was used as a tracer of macromolecular permeability. Topical application of LTB₄ (150 nM-5 M) to the hamster cheek pouch resulted in an immediate increase in adhering leukocytes in postcapillary venules and later larger venules. Leukocyte accumulation was reversible, but continued longer the higher the dose of LTB₄ used. Subsequently, a dose-dependent increase in vascular permeability was seen at postcapillary and larger venules, with a maximum 10–20 min after application; the maximum occurred later the higher the dose of LTB₄. Depletion of neutrophil granulocytes by pretreatment of the animals with antineutrophil serum obtained from immunized rabbits significantly decreased the permeability response to LTB₄, whereas the response to histamine was unaffected. These results suggest that neutrophil granulocytes play a role in LTB₄-mediated permeability increase. LTB₄ may be of importance both for the leukocyte accumulation and for the edema formation seen in inflammatory reactions.     

Mechanism of antianaphylactic action of beta-agonists in allergic inflammation of air pouch type in rats

Using an experimental model of allergic inflammation of air pouch type in rats, the mechanism of antiallergic action of beta-agonists was examined. In this model an immediate increase in vascular permeability and histamine level in the pouch fluid was observed after injecting the antigen (azobenzene arsonate-conjugated acetyl bovine serum albumin) solution into the preformed air pouch on the back of the sensitized rats. The same type of reaction was inducible by injecting anti-rat IgE into the preformed air pouch, but not IgG2a. This fact indicates that the immediate increase in vascular permeability and histamine level is an IgE-mediated anaphylactic reaction. When beta-agonists such as isoproterenol, procaterol and salbutamol were injected into the air pouch together with the antigen, the anaphylactic increase in vascular permeability was suppressed dose-dependently without concomitant decrease in histamine level in the pouch fluid. In contrast, disodium cromoglycate, an inhibitor of degranulation of mast cells, the anaphylactic vascular permeability increase was suppressed in parallel with a decrease of the histamine level. Propranolol, a beta-antagonist, counteracted the effect of beta-agonists. Serotonin-induced vascular permeability was also suppressed dose-dependently by treatment with beta-agonists. Furthermore, vascular permeability in the postanaphylactic phase of the present experimental model was also suppressed by isoproterenol. These results indicate that beta-agonists exert their antiallergic effect by inhibiting the reactivity of local vasculature to chemical mediators released from mast cells.     

Induction of neutrophil infiltration by rat chemotactic cytokine (CINC) and its inhibition by dexamethasone in rats

In vivo effects of cytokine-induced neutrophil chemotactic factor (CINC) derived from rats on neutrophil infiltration were investigated using an air-pouch-type inflammation model in rats, and effects of dexamethasone on neutrophil infiltration induced by CINC was also examined in order to gain further insight into the mechanism of antiinflammutory activity of glucocorticoids. Injection of CINC into the air pouch made on the dorsum of rats induced a marked infiltration of neutrophils into the pouch fluid but not mononuclear cells and eosinophils during a 30-min interval after the injection. Maximum effect was induced at a dose of 1.4g/pouch. Treatment with dexamethasone 3 h before the injection of CINC suppressed the neutrophil infiltration in a dose-dependent manner, but no complete inhibition was observed. CINC injection into the air pouch of rats that had been sacrificed by bleeding in order to minimize neutroph il infiltration from blood stream also stimulated neutrophil infiltration into the pouch fluid when the carcass was incubated at 37C for 30 min, but the number of infiltrated neutrophils was about 35% of CINC-induced neutrophil infiltration in intact ruts. CINC-induced neutrophil infiltration in the carcass, which is supposed to be a reflection of neutrophil migration from extravascular space in subcutaneous tissues to pouch fluid, was not inhibited by dexamethasone treatment. Therefore, the inhibition of neutrophil infiltration by dexamethasone might be due to inhibition of the extravasation of peripheral neutrophils but not due to inhibition of neutrophil chemotaxis from subcutaneous extravascular space to pouch fluid. These findings suggest that clinical effects of steroidal antiinflammatory drugs on neutrophil infiltration in inflammatory disease is partly due to inhibition of neutrophil extravasation induced by preformed neutrophil chemotactic factors in the inflammatory site.     

Changes in histamine and prostaglandins in acute inflammatory air pouch in mice

The changes in histamine and prostaglandins (PGE₂ and PGF) in carrageenan-induced acute inflammation were studied in 6 day old air pouches in mice. A significant elevation of exudate histamine was found 1 hour after the carrageenan injection. Highest vascular permeability at the site of inflammation was also found at 1 hour. Both PGE₂ and PGF showed steady increases in the pouch exudate and reached significantly higher levels at 24 hours. The present findings thus suggest that histamine is involved in the early phase of carrageenan-induced inflammation by enhancing vascular permeability. The increases in PGE₂ and PGF appear to be closely correlated with the increased exudate cell accumulation. This leads us to suggest that they are likely to be involved in the exudate cell activity rather than in enhancing the vascular permeability which was found to decrease at 4 hours after the carrageenan injection.     

Arabinogalactan- and dextran-induced ear inflammation in mice: Differential inhibition by H1-antihistamines, 5-HT-serotonin antagonists and lipoxygenase blockers

Intravenous injection of arabinogalactan or dextran together with pontamine sky-blue dye into mice increased vascular permeability and led to marked blueing of the ears. Arabinogalactan caused a rapidly progressing ear blueing (maximal coloration 20–30 min after injection). This response was suppressed by pretreating the animals with the histamine H₁-antihistamines levocabastine and loratadine. In contrast, dextran induced a slowly evolving ear inflammation (maximal coloration 60–90 min after injection), which was blocked by the 5-HT-serotonin antagonists cinanserin, metergoline and ritanserin. Furthermore, the dextran reaction was inhibited by the lipoxygenase (LO)/cyclooxygenase (CO) inhibitors BW540C, BW755C and phenidone and by the specific 5-LO inhibitor AA-861. Both arabinogalactan and dextran responses were inhibited by aprotinin, a kallikrein inhibitor, and the mixed H₁/5-HT antagonists astemizole and azatadine. The inflammogenic activity of the polysaccharides was not affected by administration of the CO inhibitors indomethacin and suprofen, the thromboxane synthetase inhibitor dazoxiben, the H₂-antihistamines cimetidine and ranitidine, the anticholinergics isopropamide or the PAF-antagonist L-652,731.These data indicate the existence of distinctive endogenous molecules that mediate the pinnal extravasation reaction to both polysaccharides: histamine for arabinogalactan, serotonin and lipoxygenase-derived arachidonic acid metabolites for dextran.     

Studies on carrageenin air pouch inflammation in the rat

Inflammation was induced in the 6-day subcutaneous air pouch of the rat by injection of carrageenin. The model was characterized in terms of exudate volume, leucocyte accumulation, granuloma, vascular permeability and protein clearance up to 7 days after injection of carrageenin. From days 2-3 rapid and reproducible changes in these responses were observed which indicated a change from polymorphonuclear (PMN) leucocyte-dominated to mononuclear (MN) leucocyte-dominated inflammation. A second injection of carrageenin on day 3 gave increases in exudate formation and PMN accumulation on day 4. Administration of carrageenin mixed with 3 day inflammatory exudate gave an increased exudate volume and decreased leucocyte accumulation at 6 h. Reduction of 6-h cellular accumulation by use of a lower dose of carrageenin or a I-day air pouch gave complete inhibition of exudate formation on day 3. In contrast, inhibition of the 6-h cell response with prednisolone had no effect on the 3-day response. Daily treatment with indomethacin gave increased PMN accumulation on day 3. Similar treatment with prednisolone additionally reduced exudate volume. Treatment on day 2 with prednisolone gave similar effects whereas indomethacin, BW755C and protease inhibitors had no effect. Administration of colchicine at this time gave inhibition of exudate volume on day 3 whereas complement depletion gave increases in volume and PMNs.     

Studies on carrageenin air pouch inflammation in the rat.

Inflammation was induced in the 6-day subcutaneous air pouch of the rat by injection of carrageenin. The model was characterized in terms of exudate volume, leucocyte accumulation, granuloma, vascular permeability and protein clearance up to 7 days after injection of carrageenin. From days 2-3 rapid and reproducible changes in these responses were observed which indicated a change from polymorphonuclear (PMN) leucocyte-dominated to mononuclear (MN) leucocyte-dominated inflammation. A second injection of carrageenin on day 3 gave increases in exudate formation and PMN accumulation on day 4. Administration of carrageenin mixed with 3 day inflammatory exudate gave an increased exudate volume and decreased leucocyte accumulation at 6 h. Reduction of 6-h cellular accumulation by use of a lower dose of carrageenin or a I-day air pouch gave complete inhibition of exudate formation on day 3. In contrast, inhibition of the 6-h cell response with prednisolone had no effect on the 3-day response. Daily treatment with indomethacin gave increased PMN accumulation on day 3. Similar treatment with prednisolone additionally reduced exudate volume. Treatment on day 2 with prednisolone gave similar effects whereas indomethacin, BW755C and protease inhibitors had no effect. Administration of colchicine at this time gave inhibition of exudate volume on day 3 whereas complement depletion gave increases in volume and PMNs.     

Quantitation of phototoxic hyperemia and permeability to protein: I. Inhibition by histamine (H1 and H2) and serotonin receptor antagonists in pig skin

Exposure of pig skin treated with a solution of anthracene to long-wave ultraviolet radiation (UVA) produced an inflammatory response which consisted of erythema and increased vascular permeability. The erythema was measured by the increase in ⁵¹Cr-RBC content of the skin; permeability of the microvasculature was determined by measurement of the accumulation of ¹²⁵I-albumin. Within the first 100 seconds (2.6 × 10³ J/M²) of irradiation, the ⁵¹Cr-RBC content of the skin increased to twice normal and remained at that level despite continued irradiation. The dermal vasculature became increasingly permeable through 1000 sec of irradiation at which time the ¹²⁵I-albumin content was ten times normal. Pretreatment of pigs with the histamine receptor antagonists pyrilamine (H₁) or cimetidine (H₂) had no effect in blocking the photobiologic increase in blood content but showed a significant inhibition of the increased permeability to ¹²⁵I-albumin. The serotonin receptor antagonist methysergide had the same permeability inhibiting effects but was about ten times more active. Thus, in the pig, the erythema occurring during the anthracene-UVA reaction is not mediated by receptors for histamine (H₁ or H₂), or by serotonin, whereas the increased vascular permeability to ¹²⁵I-albumin occurring during the same phototoxic reaction is mediated by histamine and serotonin receptors.     

Microvascular actions of histamine: synergism with leukotriene B4 and role in allergic leucocyte recruitment

Background Previous studies have shown that antihistamities provide little or no protection against the recruitment of leucocytes in allergic inflammation. Objective We wanted to examine if threshold doses of histamine can potentiate chemoattractant-induced leukocyte adhesion and if complete inhibition of histamine-induced microvascular effects is necessary to reduce allergic leucocyte recruitment. Methods The role of histamine in allergic leucocyte recruitment was examined by use of intravital microscopy of the hamster cheek pouch microcirculation. Results We found that topical administration of histamine caused a concentration-dependent increase in microvascular permeability in the cheek pouch; i.e. 0.3 μM histamine caused no detectable plasma leakage, while 1 μM and 10 μM histamine resulted in 29 ± 9.3 and 356 ± 47 leakage sites/cm² cheek pouch area, respectively. The percentage of postcapillary venules with more than five adherent leucocytes (an index of early leucocyte recruitment) was 1.1 ± 0.51% in the control situation, and did not increase significantly after stimulation with histamine alone (0.3–10μM) or with 1 nM ieukotriene B₄ (LTB₄). On the other hand, coapplication of 10μM histamine and 1 nM LTB₄ increased leucocyte adhesion 24-fold. In fact, the 10 times lower dose of histamine (1 μM) together with 1 nM LTB₄ increased leucocyte adhesion to a similar extent (20 fold). The increase in vascular permeabihty evoked by exogenous 10μM histamine (with or without LTB₄), or by histamine released from activated mast cells (antigen challenge), was completely reversed by local pretreatment with the H₁-receptor antagonist mepyramine. This mepyramine treatment also abohshed the enhanced leucocyte adhesion in response to coapplication of histamine and LTB₄. Moreover, mepyramine, which had no effect on leucocyte recruitment evoked by 3 nM LTB₄per se, reduced antigen-induced recruitment of leucocytes to the extravascular tissue by 79.5 ± 14.8%. Conclusion We conclude that threshold concentrations of histamine can strikingly potentiate chemoattractant-induced leucocyte responses, and that in order to reduce allergic leucocyte recruitment it may be necessary to use antihistamines in doses high enough to abolish the microvascular actions of histamine.     

Possible role for platelet-activating factor in neutrophil infiltration in allergic inflammation in rats

Allergic inflammation was induced by injecting an antigen solution into an air pouch made on the dorsum of immunized rats with the antigen azobenzene-arsonate-conjugated acetyl bovine serum albumin. In this model, leukocyte infiltration into the pouch fluid was prominent 4-8 h after the antigen challenge. Most of the infiltrated leukocytes were neutrophils. Administration of the platelet-activating factor (PAF) antagonists such as CV-3988 and L-652,731 into the air pouch 15 min before and at the time of the antigen challenge failed to suppress leukocyte infiltration at 8 h. However, when the PAF antagonist was injected into an air pouch 4 h after the antigen challenge, neutrophil infiltration at 8 h was suppressed in a dose-dependent manner. Combined treatment with the 5-lipoxygenase inhibitor AA861 and the PAF antagonist did not potentiate the effect of the PAF antagonist, suggesting that participation of leukotriene B4 in neutrophil infiltration in this model is negligible. Eosinophil infiltration was very weak at 8 h, and the PAF antagonist showed no significant effect. At 8 h, the PAF level in the serum of the immunized rats was significantly higher than that of the nonimmunized rats. Intravenous administration of the PAF antagonist 15 min before the antigen challenge suppressed leukocyte infiltration more effectively than local administration into the pouch. These results indicate that PAF plays a significant role in neutrophil infiltration in allergic inflammation.     

Changes in corticosterone levels under different degrees of acute inflammation in mice

Carrageenan of different concentrations was injected into the 6-day-old air pouch in mice. It was found that 12 mg carrageenan caused a significant increase of plasma and exudate corticosterone levels at 24 h, while 1 and 3 mg carrageenan could only induce a significant increase of exudate corticosterone at 4 h. Elevation of corticosterone in both plasma and inflammatory exudate appeared to be correlated, suggesting that the exudate corticosterone was derived from the blood circulation. Injection of exogenous histamine and PGE₂ into the air pouch induced a significant increase in exudate levels of corticosterone. However, plasma corticosterone increased significantly only after histamine administration, although a slight increase was observed in those injected with PGE₂. These findings thus suggest that endogenous histamine and PGE₂ which are released during carrageenan-induced acute inflammation, as shown in our previous work, might be responsible for the increase of corticosterone in both plasma and inflammatory exudate.     

Further studies on the anti-inflammatory effect of insulin

Experiments performed on rats showed that insulin, when applied i.v. or s.c., inhibited the foot edema induced by carrageenin, thermic effect of 45.7° C, compound 48/80 and 5-HT, but moderately increased the paw swelling evoked by kallikrein, a kinin-forming enzyme. The increased vascular permeability elicited by intradermal injection of histamine, 5-HT, bradykinin, PGE₁, carrageenin and compound 48/80 was also suppressed. The anti-inflammatory effect was not significantly altered by propranolol and adrenalectomy on the thermal and carrageenin edema, it was variably inhibited on the skin test, and was completely abolished on the paw swelling induced by 5-HT and compound 48/80. Since insulin had little or no effect on the vascular response when given topically together with the vasoactive agents, its complex effect on the acute inflammation appears to be brought about via indirect mechanisms.     

Increase in Histamine Production by Inflammatory Exudate in the Chronic Phase of Allergic Inflammation in Rats

In the air pouch-type allergic inflammation in rats, we reported that a sustained histamine production in the late phase is induced by a cytokine-like factor, named histamine-production-increasing factor (HPIF) (1). Recently, we found another type of histamine-production-increasing factor in the pouch fluid at the chronic phase of air pouch-type allergic inflammation. Although it did not increase histamine production by itself, it enhanced the HPIF-induced histamine production by rat bone marrow cells. It also increased GM-CSF-induced histamine production. The activity of this factor increased time-dependently from 3 to 7 days after the antigen challenge. Injection of the 5 day pouch fluid sample containing this factor into the pouch 4 h after the antigen challenge increased histamine contents in the pouch fluid at 24 h, indicating that this factor enhances HPIF-induced histamine production in vivo. Biochemical analysis of the 5 day pouch fluid sample indicated that this factor is a heat-labile and trypsin-sensitive protein of which pI value and molecular weight are 7–8 and about 100 kDa, respectively.     

Central histaminergic stimulation of hyperlipemic response in rats under stress

In rats subjected to a mild stress of immobilization histamine, the H₁-receptor agonist 2-pyridylethylamine (PEA), and the H₂-receptor agonists 4-methyl histamine (4-MeHA) and impromidine administered intracerebroventricularly (i.c.v.) 1 h prior to stress, intensified the stress-induced increase in serum free fatty acid (FFA) levels. Impromidine was far more potent than histamine and its agonists in increasing hyperlipemia in stressed rats. The hyperlipemic response to histamine was abolished by i.c.v. pretreatment of rats with mepyramine, a H₁-receptor antagonist, but was unchanged in rats pretreated with cimetidine or metiamide, H₂-receptor antagonists. The increase in serum FFA levels induced in stressed rats by PEA was abolished by mepyramine but the hyperlipemic responses to 4-MeHA and impromidine were not antagonized by cimetidine.These results suggest that central H₁-receptor mediate the histamine-stimulated hyperlipemic response in stressed rats.     

Effect of histamine receptor antagonists on functional hyperaemia in skeletal muscles in cats

Conclusions The results show that H₁, and H₂ receptor antagonists, used together, result in a diminishing vasodilatator mechanism which contributes to vasodilatation accompanying and following functional activity. They suggest that histamine may be involved in the local vascular response to exercise in the vascular bed of skeletal muscles of the cat.     

Histamine H1- and H2-receptor involvement in eosinophil infiltration and the microvascular changes associated with cutanneous anaphylaxis

Several substances alter eosinophil motility, but the relative importance of these putative mediators in immediate hypersensitivity remains unclear. The present study has reinvestigated the role of histamine in type I allergic eosinophil infiltration, and the temporally associated microvascular events, by examining the effect of H₁-and H₂-receptor antagonist pretreatment. A combination of cimetidine and pyrilamine significantly reduced eosinophil accumulation, whereas neither antagonist alone was effective. Similarly, cutaneous hyperemia, measured indirectly as ear surface temperature, was reduced only by the cimetidine-pyrilamine combination. Pyrilamine partially attenuated the increase in microvascular permeability, but the addition of cimetidine provided no further reduction.It appears that histamine participates significantly in mediating both the microvascular changes and the eosinophil infiltration evoked by cutaneous anaphylaxis. The histaminergic component of increased microvascular permeability appears to be an H₁-receptor mediated phenomenon. However, blockade of both H₁-and H₂-receptor subtypes is required to inhibit the hyperemia and eosinophil infiltration responses.     

Quantitative physiological and morphological aspects of microvascular permeability changes induced by histamine and inhibited by terbutaline

Histamine induced changes in microvascular permeability to macromolecules were studied in two models, the hamster cheek pouch and the canine forelimb preparation. Fluoresceinlabelled dextran FITC-dextran. M_w=70000 and 150000 was used as tracer of permeability increases. Studies in the hamster showed that morphological changes as indicated by intravital microscopy of FITC-dextran leakage at postcapillary venules were accompanied by an increased concentration of FITC-dextran in the superfusion solution for the cheek pouch. Following histamine challenge during 4 min the cheek pouch microcirculation was normalized 30 min later both with regard to number of leakage sites and the concentration of FITC-dextran in the superfusion solution. Pretreatment of the cheek pouch with terbutaline (3.6 × 10^-6 M) for 1 h resulted in total inhibition of both morphologically and fluorometrically recorded changes from continuous histamine superfusion. When terbutaline and histamine were given at the same time, there was a significant inhibition of the histamine induced increase in the number of leakage sites to 10% of the pre-terbutaline response. The fluorometrically detected efflux of FITC-dextran was 65% of the preterbutaline value. The dynamics of interendothelial cell gap formation and closure was also studied in the dog. Intra-arterial infusions of histamine into forelimbs perfused at a constant pump controlled flow rate produced decreases in perfusion pressure, increases in lymph flow, lymph protein concentration, total protein transport and weight. FITC-dextran was given as an i. v. injection 30 min before and 0, 30 and 60 min after the start of a 90 min arterial infusion of histamine into the brachial artery of the forelimb. Lymph (C_L) and plasma (C_p) concentrations of FITC-dextran were determined and C_L/C_p-ratios showed that the permeability increasing effect lasted less than 30 min although histamine was given for further 60 min. The results of both hamster and forelimb experiments suggest that leakage sites or interendothelial cell gaps in postcapillary venules are closed within 30 min from the first exposure to histamine, probably within 15 min and that there is little indication of any remaining increase in macromolecular permeability via any other route (vesicular) following closure of gaps.