6-trans-leukotriene B4 is a neutrophil chemotaxin in the guinea pig dermis.

The products of the 5- and 12-lipoxygenase (5-LO, 12-LO) pathways of arachidonic acid metabolism are implicated as proinflammatory mediators in a number of disease states. 12(R)-hydroxyeicosatetraenoic acid [12(R)-HETE] is present in large quantities in human psoriatic lesional skin and can be further metabolized by 5-LO to 5(S), 12(R)-dihydroxy-(6E,8Z,10E,14Z)-eicosatetraenoic acid (6-trans-LTB4). Furthermore, leukotriene B4 (LTB4) and the sulfidopeptide leukotrienes (LTC4, LTD4) can be transformed to 6-trans-LTB4. When injected into the guinea pig dermis, 6-trans-LTB4 (1.0, 10.0, 20.0 micrograms/intradermal site) caused a significant (P less than 0.02) infiltration of polymorphonuclear leukocytes (PMN) at 4 hr as assessed by histology and the levels of the PMN marker enzyme myeloperoxidase. 6-trans-LTB4 is a more potent PMN chemoattractant than 12(R)-HETE in the guinea pig dermis but is far less potent than LTB4. Pharmacological interdiction of leukotriene production or receptor binding should take into account the proinflammatory activity of 6-trans-LTB4.     

Leukotriene B4-induced granulocyte trafficking in guinea pig dermis

Leukotriene B₄ (LTB₄) is a proinflammatory product of arachidonic acid metabolism that has teen implicated as a mediator in a number of inflammatory diseases. When injected intradermally into the guinea pig, LTB₄ elicits a dose-dependent migration (chemotaxis) of neutrophils (PMNs) into the injection sites as assessed by the presence of a neutrophil marker enzyme myeloperoxidase. SC-41930 7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxy]-3,4-dihydro-8-propyl-2H-1 -benzopyran-2-carboxylic acid, a first-generation LTB₄ receptor antagonist inhibitedthe chemotactic actions of LTB₄ when coadministered into the dermal site and when given orally with ED₅₀ values of 340 ng and 1.7 mg/kg, respectively. The secondgeneration LTB₄ receptor antagonists SC-50605 7-[3-[2(cyclopropylmethyl)-3-methoxy-4-(4-thiazolyl)phenoxy] propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid and SC-51146 7-[3-[2(cyclopropylmethyl)-3-methoxy-4-[(methylamino)carbonyl]phenoxy]propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran2-propanoic acid inhibited LTB₄-induced chemotaxis when coadministered with ED₅₀ values of 70 ng and 32 ng, respectively, and when given intragastrically with ED₅₀ values of 0.10 and 0.09 mg/kg, respectively. SC-41930, SC-50605, and SC-51146 had oral durations of action of 5.5, 15, and 21 h, respectively. These potent, LTB₄ receptor antagonists may well have application in the medical management of disease states such as asthma, rheumatoid arthritis, inflammatory bowel disease, contact dermatitis, and psoriasis, where LTB₄ is implicated as an inflammatory mediator.     

Mechanisms of leukotriene-induced contractions of guinea pig airways: leukotriene C4 has a potent direct action whereas leukotriene B4 acts indirectly

The leukotrienes (LT's) are a group of arachidonic acid derivatives implicated as mediators of allergic bronchoconstriction and acute inflammation. Tracheal spirals and strips of lung parenchyma from guinea pigs were used under non-flow conditions to characterize the contractions caused by LTA4, LTB4 and LTC4. Cumulative administrations of leukotrienes desensitized the lung strip, whereas non-cumulative dose-response relationships for the leukotrienes and histamine were reasonably parallel. Half maximal contractions of the lung strips were obtained at a final bath concentration of 1 nM for LTC4 and 300 nM for LTA4 or LTB4, as compared with 6 000 nM for histamine. In the trachea, LTC4 was approximately 100 times more potent than LTA4 and histamine. Leukotrienes B4 and C4, but not acetylcholine or histamine, elicited release of the bronchoconstrictive thromboxane A2 from the lung under non-flow conditions. Indomethacin blocked the contractile response to LTB4, whereas the contractile effect of LTC4 remained unaltered. The beta-adrenoceptor agonist isoproterenol and the LTC4 antagonist FPL 55712 attenuated the contraction, but not the release of thromboxane A2, induced by LTC4. Changing to a perifusion technique rendered the lung strips less sensitive to the direct action of LTC4, and released thromboxane A2 now contributed significantly to the contractile response. In addition, the perifusion experiments indicated that LTB4 released histamine as well. We conclude that the chemoattractant LTB4 is an indirectly acting bronchoconstrictor, whereas the slow reacting substance LTC4 contracts the airway muscle by a predominantly direct mechanism. The exquisite bronchoconstrictive activity of LTC4 may be unrelated to its ability to induce formation of thromboxane A2.     

Anaphylatoxin C3a but not C3a(desArg) is a chemotaxin for the mouse macrophage cell line J774

Varying results have been published regarding the functional reactivity of different cell types, including human monocytes, to the anaphylatoxin C3a and its degradation product C3a(desArg). To further delineate the functions of C3a and C3a(desArg) on this cell type we used the murine macrophage (Mø) cell line J774A.1 which is known to respond to the anaphylatoxin C5a. J774 cells specifically bound fluorescein isothiocyanate-labeled recombinant human C3a (rC3a). The cells migrated along rC3a concentration gradients in a dose-dependent manner with an optimal concentration of about 3 nM (rC5a: 7 nM) and a half-maximal effective concentration (EC₅₀) of about 1.2 nM (rC5a: 2 nM). The degradation product rC3a(desArg) was devoid of chemotactic activity. mRNA for the recently cloned G protein-coupled mouse high-affinity C3a receptor (C3aR) was detected in J774 cells, suggesting that this receptor represents the binding site for C3a on J774 Mø. In support of the specific nature of C3a-stimulated cellular mobility, RBL-2H3 transfectants expressing the human C3aR were also shown to migrate along gradients of rC3a (optimal concentration about 8 nM; EC₅₀ about 3.5 nM) whereas rC3a(desArg) was again inactive. In summary, our findings demonstrate for the first time a specific, receptor-mediated chemoattraction of cells of the monocytic lineage to the anaphylatoxin C3a which may contribute to the accumulation of Mø at sites of inflammation.     

Immune response to a thymus-independent antigen (DNP-Ficoll) in the guinea pig

Guinea pigs immunized with DNP30-Ficoll produced IgM antibody only. No IgG1, IgG2, IgE antibodies or delayed hypersensitivity were detected in these animals. However, Arthus reactions, induced by the hapten coupled to a foreign carrier or the whole antigen, were found. The time course of the IgM response was limited and the response to reinjection of the antigen reduced. Cyclophosphamide (CY), given 3 days before primary immunization, prolonged the IgM response. Given on the day of immunization or 3 days after CY reduced this response. CY given on days +3 or +7 after primary immunization completely suppressed the response to antigen reinjected 42 days later. Arthus reactions were totally suppressed by CY given on the day of immunization, or 3 or 7 days later.     

Experimental neonatal syphilis in a susceptible (C4D) and a resistant (Albany) strain of guinea pig.

Despite similar levels of natural antibodies and treponemicidal activity, 83% of fourth complement component-deficient (C4D) mother guinea pigs developed ulcerative lesions to a challenge of 5 x 10(7) Treponema pallidum, whereas 75% of offspring 1 to 5 days old were temporarily (2-3 months) resistant to development of dermal lesions. In contrast, only 17% of Albany-strain mothers developed small papular lesions, while 68% of 1- to 5-day-old newborns developed large papular or ulcerative lesions within 9-15 days postinfection. These findings, together with the late development of both dermal lesions and antibodies in C4D neonates, preclude the concept of an antibody-associated natural resistance. T. pallidum infection in either C4D or Albany neonates was not associated with depletion of any particular cell population in lymphoid tissue. However, marked age- and strain-dependent histologic differences were noted. Histologic examination of lymph nodes and spleens from 17-day-old and 3- to 4-month-old animals showed that maturation of lymphoid tissues in C4D animals lagged behind the Albany strain at either age. Moreover, 75% of C4D newborns contained significantly higher levels of immunomodulatory alpha 1 fetoprotein than Albany neonates. The possibility that differences in susceptibility to T. pallidum infection between C4D and Albany guinea pigs as neonates and again as adults is the result of genetically associated changes in immunologic recognition is discussed.     

Effect of the leukotriene B4 receptor antagonist,SC-41930, on experimental allergic encephalomyelitis (EAE) in the guinea pig

The accepted model for the human demyelinating disease, multiple sclerosis (MS), is experimental allergic encephalomyelitis (EAE). We assessed the ability of SC-41930(7-[3(4-acetyl-3-methoxy-2-propylphenoxy)-propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxyl acid), to modulate the symptoms of acute EAE generated in guinea pigs. Animals were pretreated with SC-41930 (20 mg/kg, i.p.) for two days followed by thrice-weekly maintenance. At day 52, a significant number of the SC-41930-treated animals were alive as compared to EAE alone. Control animals had an increase in body weight while EAE animals lost over 20% (p<0.5) of their body weight by day 18. SC-41930-treatment significantly reduced, but did not completely inhibit the cachectic response. The results indirectly implicate LTB₄ in the pathogenesis of EAE. Agents that modify this model be useful in the treatment of human MS.     

Inhibition of IKs in guinea pig cardiac myocytes and guinea pig IsK channels by the chromanol 293B

The chromanol derivative 293B was previously shown to inhibit a cAMP regulated K⁺ conductance in rat colon crypts. Subsequent studies on cloned K⁺ channels from the rat demonstrated that 293B blocks specifically I_sK channels expressed in Xenopus oocytes, but does not affect the delayed and inward rectifier Kv1.1 and Kir2.1, respectively. In the present study, the specificity of 293B for the cardiac K⁺ conductances I_Ks and I_Kr, and for the cloned guinea pig I_sK channel and the human HERG channel, which underly I_Ks and I_Kr, respectively, was analyzed. 293B inhibited both the slowly activating K⁺ conductance I_Ks in cardiac myocytes and guinea pig I_sK channels expressed in Xenopus oocytes with a similar IC₅₀ (2-6 μmol/1). In contrast, high concentrations of 293B had only a negligible effect on the more rapid activating I_Kr. Similarly, 293B exerted no effect on HERG channels expressed in Xenopus oocytes. In summary, 293B appears to be a rather specific inhibitor of I_Ks and the underlying I_sK channels.     

ACTH(4-10) accelerates ocular motor recovery in the guinea pig following vestibular deafferentation

Neuropeptide hormones such as adrenocorticotropic hormone, fragment 4-10 (ACTH(4-10], have been shown to facilitate various kinds of CNS plasticity, including recovery from deafferentation of the inner ear (vestibular compensation). The purpose of the present experiment was to determine whether the rapid compensation of spontaneous nystagmus (SN), which occurs over 2-3 days post-unilateral labyrinthectomy (UL) in the guinea pig, could be accelerated by administration of ACTH(4-10). Because of the short half-life of ACTH(4-10), injections of 200 micrograms/kg i.m. were given every 4 h for 48 h post-UL, and SN was measured every 2 h for 52 h post-UL. The results were compared with SN measurements from guinea pigs which received saline injections of the same volume, at the same times. ACTH(4-10) injections were found to significantly accelerate the rate of compensation of SN following UL. This result suggests that ACTH(4-10) may be useful in facilitating compensation when the symptoms of UL are most severe, during the first 2-3 days post-UL.     

Effect of a leukotriene B4 receptor antagonist on leukotriene B4-induced neutrophil chemotaxis in cavine dermis

Leukotriene B₄ (LTB₄) is a proinflammatory product of arachidonic acid metabolism that has been implicated as a mediator in a number of inflammatory diseases. When injected intradermally into the cavine, LTB₄ elicits a dose-dependent immigration (chemotaxis) of neutrophils (PMNs) into the injection sites as assessed by the presence of a neutrophil marker enzyme myeloperoxidase. SC-41930 {7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxy]-3,4-dihydro-8-propyl-2H-l-benzopyran-2-carboxylic acid, a potent LTB₄ receptor antagonist inhibited the chemotactic actions of LTB₄ when coadministered into the dermal site and when given intravenously or orally with ED₅₀ values of 200 ng, 0.5 mg/kg, and 0.6 mg/ kg respectively. This compound may well have application in disease states, such as inflammatory bowel disease and psoriasis, where LTB₄ is implicated as a proinflammatory mediator.     

Thermosensitivity of preoptic neurones in a hibernator (golden hamster) and a non-hibernator (guinea pig)

Summary Thermosensitivity of preoptic units was studied at hypothalamic temperatures (T _hy) ranging from 8–43°C in golden hamsters in a non-hibernating state as well as in guinea pigs. In golden hamsters 2 types of thermoresponsive preoptic neurones were found: 1. Neurones sensitive toT _hy ranging from 10–42°C with an exponential characteristic and very high spontaneous firing rates (29–59 imp/s) atT _hy 36–37°C. 2. Neurones with a bell-shaped temperature-firing rate characteristic, a negative temperature coefficient atT _hy 40–30°C, a maximal activity atT _hy 20–30°C and a positive temperature coefficient (+0.8 to +4 imp/s·°C) even atT _hy close to 10°C. In guinea pigs thermoresponsive preoptic units became inactive or insensitive to thermal stimulation as soon asT _hy fell below 30°C. These results suggest that in hibernators central nervous structures involved in temperature regulation are adapted to maintain their function over the wide range of core temperatures which occur during the different phases of hibernation.Supported by the Deutsche Forschungsgemeinschaft, Project B1. SFB 122.     

Gastric HCO3--secretion in the guinea pig.

Measurement of gastric intraluminal PCO2 and pH in the anesthetized guinea pig enabled simultaneous determination of total H+ and HCO3- gastric secretions. There was quantitative agreement between the release of CO2 and decrease in HCO3- after intragastric instillation of HCl. The basal rate of HCO3- secretion (approximately 40 mueq-h-1) was, in most cases, smaller than spontaneous H+ secretion, but gastric net secretory output was alkaline (HCO3- greater than H+) after inhibition of acid secretion with histamine H2-receptor antagonists (cimetidine 20 mg-kg-1 or metiamide 35 mg-kg-1). Carbachol (1-2 microgram-kg-1) stimulated secretion of both HCO3- and H+; only the latter response was sensitive to the histamine antagonists. Atropin (100 microgram-kg-1) blocked stimulation of HCO3- secretion but did not affect the basal output of HCO3-. An increase in HCO3- secretion was associated with an equivalent increase in net Na+ influx and an increase in the net influx of Cl- with H+ plus K+. Intragastric neutralization of H+ by HCO3- is likely to occur at the mucosal surface and may protect the mucosa from the damaging effects of intraluminal acid.     

Pinealectomy and two-way avoidance in the guinea pig (Cavia porcellus)

The effects of pinealectomy on two-way avoidance learning in the guinea pig were investigated. Results showed no difference between pinealectomized and normal guinea pigs on any of the performance measures taken. This suggests that blindness-induced facilitation of two-way avoidance in guinea pigs is not a result of the hormonal changes in the pineal known to accompany variations in visual input.     

Ultrastructure of basophilic leukocytes and mast cells in normal and cutaneous basophil hypersensitivity-reacted guinea pig dermis.

Basophilic leukocytes and mast cells in guinea pig dermis in normal and cutaneous basophil hypersensitivity (CBH) reaction were examined by light and electron microscopy. Basophils were rare in the normal dermis and predominantly revealed in the CBH-reacted skin. Some infiltrating basophils of the reacted displayed an immediate attachment to mast cells. Cytoplasmic continuities were partially seen between them. They were most plentiful at 48 h after phytohemagglutinin injection and decreased in number thereafter. The basophils contained three types of granule. The vast majority of granules were Type I granules, basophil-specific granules. Type II granules were less frequently encountered and resembled a mast cell granule in the fine structure. Type III granules were scant and small-cored vesicles.     

Vestibular and auditory cortical projection in the guinea pig (cavia porcellus)

Summary Isolated electrical stimulation of the vestibular nerve in the guineapig yielded surface positive evoked potentials within the rostral portion of the SI forelimb field. The locus of negative field potential reversal in deeper cortical layers was small. The vestibular field is distinct from those of the auditory and facial nerves. Comparative aspects of vestibular cortical location are discussed. The auditory field corresponds with that of other rodents.Part of this study is to be published as a Congress abstract (Ödkvist et al., 1972).This research was supported by Medical Research Council grant # 3311, and the Hartford Foundation.     

Interleukin-8 (IL-8) is a major neutrophil chemotaxin from human alveolar macrophages stimulated with staphylococcal enterotoxin A (SEA)

Since Staphylococcus aureus is an important human pathogen, and infection of the lungs in characterized by neutrophil infiltration we studied the role of a staphylococcal toxin, enterotoxin A (SEA) on the synthesis and secretion of IL-8 by human alveolar macrophages. As SEA concentration was increased, the IL-8 accumulation in the macrophage conditioned medium increased. The concentration of mRNA encoding IL-8 was also elevated in the macrophage in response to increases in SEA concentration. Although the monocytic cell line U937 was able to respond to SEA and secrete IL-8, treatment with PMA prior to SEA stimulation increased the IL-8 accumulation around fifty fold indicating that maturation of the undifferentiated cell to a more macrophage-like cell facilitated IL-8 accumulation. Stimulating human alveolar macrophages with high concentrations of SEA caused an increase in IL-1 accumulation. However, when the cells were incubated with SEA in the presence of IL-1 receptor antagonist, there was no decrease in IL-8 accumulation. Addition of a neutralizing anti-IL-8 monoclonal antibody to the culture medium of SEA-stimulated macrophages significantly reduced the neutrophil chemotactic activity of the medium. These studies showed that IL-8 is a major neutrophil chemotaxin from human alveolar macrophages stimulated with SEA.accepted by I. Ahnfelt-Rønne