Identification of an immune-responsive mesolimbocortical serotonergic system: potential role in regulation of emotional behavior.

Peripheral immune activation can have profound physiological and behavioral effects including induction of fever and sickness behavior. One mechanism through which immune activation or immunomodulation may affect physiology and behavior is via actions on brainstem neuromodulatory systems, such as serotonergic systems. We have found that peripheral immune activation with antigens derived from the nonpathogenic, saprophytic bacterium, Mycobacterium vaccae, activated a specific subset of serotonergic neurons in the interfascicular part of the dorsal raphe nucleus (DRI) of mice, as measured by quantification of c-Fos expression following intratracheal (12 h) or s.c. (6 h) administration of heat-killed, ultrasonically disrupted M. vaccae, or heat-killed, intact M. vaccae, respectively. These effects were apparent after immune activation by M. vaccae or its components but not by ovalbumin, which induces a qualitatively different immune response. The effects of immune activation were associated with increases in serotonin metabolism within the ventromedial prefrontal cortex, consistent with an effect of immune activation on mesolimbocortical serotonergic systems. The effects of M. vaccae administration on serotonergic systems were temporally associated with reductions in immobility in the forced swim test, consistent with the hypothesis that the stimulation of mesolimbocortical serotonergic systems by peripheral immune activation alters stress-related emotional behavior. These findings suggest that the immune-responsive subpopulation of serotonergic neurons in the DRI is likely to play an important role in the neural mechanisms underlying regulation of the physiological and pathophysiological responses to both acute and chronic immune activation, including regulation of mood during health and disease states. Together with previous studies, these findings also raise the possibility that immune stimulation activates a functionally and anatomically distinct subset of serotonergic neurons, different from the subset of serotonergic neurons activated by anxiogenic stimuli or uncontrollable stressors. Consequently, selective activation of specific subsets of serotonergic neurons may have distinct behavioral outcomes.

Acute pro-inflammatory actions of endothelin-1 in the guinea-pig lung: involvement of ETA and ETB receptors.

1. Although recent observations suggest that endothelin-1 (ET-1) may play a role in the pathogenesis of asthma, to date little is known about the effects of ET-1 on parameters other than bronchoconstriction. The objectives of the present experiments were to study whether intravenously administered ET-1 could exert pro-inflammatory actions in the guinea-pig lung and to assess the involvement of endothelin ETA and ETB receptors in these events by using the ETA receptor-selective antagonist, FR 139317, the novel ETA/ETB receptor antagonist, bosentan and the ETB receptor-selective agonist, IRL 1620. 2. Bolus i.v. injection of ET-1 (0.1-1 nmol kg-1) to anaesthetized guinea-pigs evoked dose-dependent increases in mean arterial blood pressure which lasted for 6-12 min. This was accompanied by a dose-dependent haemoconcentration (8-15% plasma volume losses) and increases (up to 546%) in albumin extravasation in the trachea, upper and lower bronchi, but not in the pulmonary parenchyma. Qualitatively similar changes were observed following i.v. injection of the ETB receptor agonist, IRL 1620 (0.3 and 1 nmol kg-1), although IRL 1620 appeared to be about 3 times less potent than ET-1. The ETA receptor-selective antagonist, FR 139317 (2.5 mg kg-1) inhibited the ET-1 (1 nmol kg-1)-induced pressor response, haemoconcentration and albumin extravasation by 75, 77 and 60-70%, respectively, whereas it did not attenuate IRL 1620 (1 nmol kg-1)-induced changes. The ETA/ETB receptor antagonist, bosentan (10 mg kg-1) almost completely inhibited the pressor, haemoconcentration and permeability effects of both ET-1 and IRL 1620. 3. ET-1, but not IRL 1620 (0.1-1 nmol kg-1), produced a dose-dependent neutropenia with relative lymphocytosis and monocytosis, but did not induce influx of neutrophil granulocytes into pulmonary tissues or the bronchoalveolar space. ET-1 (1 nmol kg-1)-induced neutropenia was prevented by pretreatment of the animals with FR 139317 (2.5 mg kg-1), bosentan (10 mg kg-1) or adrenaline (90 nmol kg-1), indicating that ET-1 caused intravascular sequestration of neutrophil granulocytes. 4. ET-1 or IRL 1620 (10(-10)-10(-6) M) alone did not activate alveolar macrophages in vitro, whereas at a concentration of 10(-8) M, ET-1, but not IRL 1620, markedly potentiated superoxide production in response to f-Met-Leu-Phe (10(-9)-10(-7) M) and platelet-activating factor (PAF, 10(-9)-10(-7) M), but not to phorbol 12-myristate 13-acetate (10(-9) M). ET-1 did not affect f-Met-Leu-Phe- or PAF-induced increases in intracellular free calcium concentration. This potentiating effect of ET-1 was abolished by FR 139317(1.5 X 10-7 M).5. We conclude that, in addition to evoking airway contractions, ET-1 exerts pro-inflammatory actions via activation of the ETA and to a lesser extent the ETB receptors, and therefore, might contribute to the airway inflammation present in asthma. These findings also suggest the therapeutic potential of ETA/ETB receptor and perhaps ETA receptor-selective antagonists in this disease.

5-Lipoxygenase and endotoxin-induced microvascular albumin exchanges and leucocyte recruitment in guinea-pig lungs.

The interference of the 5-lipoxygenase inhibitor, BW B70C ((E)-N-(3-[3-(4-fluorophenoxy)phenyl]-1(R,S)-methyl prop-2-enyl)-N-hydroxyurea), with Escherichia coli lipopolysaccharide (endotoxin)-induced lung leucocyte sequestration and microvascular albumin exchanges was evaluated in the anaesthetised guinea-pig using radioactive tracers, in parallel to the effects on cell counts in the broncho-alveolar lavage fluid, blood tumour necrosis factor (TNF-alpha) content, secretion of phospholipase A2 and synthesis of leukotriene C4 by alveolar macrophages. Intravenous injections of 0.1 or 1 mg/kg endotoxin induced lung leucocyte sequestration but only the higher dose induced an increase in albumin microvascular exchanges and the infiltration of leucocytes towards the airway lumen. Leukotriene B4, a potential mediator of the 5-lipoxygenase-dependent endotoxin effects, induced a rapid and transient lung leucocyte sequestration and leucopenia associated with a more progressive increase in microvascular exchanges. The 5-lipoxygenase inhibitor, BW B70C, injected i.p. (30 mg/kg) prevented leukotriene C4 synthesis by alveolar macrophages and reduced leucocyte migration to the airways lumen as well as albumin microvascular leakage but did not affect the endotoxin-induced increase in the blood level of TNF-alpha and of secreted phospholipase A2. However, BW B70C failed to modify vascular leucocyte margination induced by 1 mg/kg endotoxin, suggesting that, apart from a role of 5-lipoxygenase, alternative pathways operate in response to endotoxin in guinea-pig.

Communicating bronchopulmonary foregut malformations in the adriamycin-induced rat model of oesophageal atresia.

BACKGROUND: Communicating bronchopulmonary foregut malformations (CBPFM) are rare abnormalities of the development of the primitive foregut that result in an abnormal communication between the upper gastrointestinal tract and pulmonary tissue. They usually occur in isolation, but sometimes are seen in association with oesophageal atresia (OA). METHODS: Communicating bronchopulmonary foregut malformations were induced in the offspring of pregnant rats by intraperitoneal injection of Adriamycin (Delta West Pty Ltd, Bentley, Western Australia, Australia). Fetuses harvested by caesarean section were fixed in 10% formalin, transversely sectioned and stained with haematoxylin and eosin. Serial examination of the slides allowed three-dimensional reconstruction of the anatomy of the pulmonary system and the oesophagus. RESULTS: Communicating bronchopulmonary foregut malformations occurred in nine (30%) of fetuses with OA. Three types of CBPFM were produced: an isolated pulmonary structure (accessory lung) attached to the lower oesophagus via a patent bronchus (6 fetuses); an anomalous bronchus from the lower oesophagus to the lower part of the left lung (two fetuses); and atresia of the trachea (one fetus). CONCLUSIONS: These observations are consistent with the assertion that CBPFM and OA are variations of a spectrum of abnormalities and may have a similar aetiology. In the rat model it would appear that Adriamycin interferes with the timing and progression of lung bud differentiation at a time when the primitive foregut is developing rapidly. Ultimately, this model may shed light on the embryogenesis of both anomalies.

Interleukin-2 induces early multisystem organ edema mediated by neutrophils.

Interleukin-2 (IL-2), an agent known to activate neutrophils (PMN) with thromboxane (Tx)B2 release, produces pulmonary edema within 6 hours of intravenous infusion. This study tests the role of PMN in mediating the edema. Anesthetized rats received 10(6)U recombinant human IL-2 (n = 15) or vehicle (n = 14) as a constant intravenous infusion during a period of 1 hour. At this time there was leukopenia 3.63 +/- 0.43 (x10(3)/mm3) relative to vehicle-infused control rats 6.12 +/- 0.86 and a decline in PMN, 2.19 +/- 0.14 relative to control value of 3.33 +/- 0.05 (both p less than 0.05). After 6 hours edema, as measured by increase in the wet to dry weight (W/d) ratio, was present in the lungs (4.93 +/- 0.20 relative to control 4.06 +/- 0.10), heart (4.09 +/- 0.11 versus 3.76 +/- 0.08), liver (3.50 +/- 0.10 versus 3.18 +/- 0.10), and kidney (4.25 +/- 0.07 versus 4.00 +/- 0.07) (all p less than 0.05). There was increased lung permeability demonstrated by bronchoalveolar lavage fluid protein concentration of 1970 +/- 210 micrograms/mL relative to control 460 +/- 90 micrograms/mL (p less than 0.05). Interleukin-2 resulted in lung PMN sequestration of 53 +/- 7 PMN/10 high-power fields (HPF) relative to 23 +/- 2 PMN/10 HPF in controls (p less than 0.05) and increased plasma TxB2 levels to 1290 +/- 245 pg/mL relative to control 481 +/- 93 pg/mL (p less than 0.05). Pretreatment of other rats (n = 8) with selective anti-rat neutrophil antiserum 18 hours before the experiment led to a peripheral PMN count 10% of baseline and prevented edema in the lungs (W/d ratio 4.20 +/- 0.16) and heart (3.67 +/- 0.07) (both p less than 0.05) but not liver or kidney. Protein in lung lavage was reduced to 760 +/- 220 micrograms/mL (p less than 0.05). The protection afforded by leukopenia was associated with lack of PMN sequestration and prevention of the increase in plasma Tx levels (484 +/- 120 pg/mL, p less than 0.05). These data indicate that the rapid induction of lung and heart edema with a 1-hour infusion of IL-2 in the rat is mediated, in large part, by activated PMNs.

The importance of aprotinin and pentoxifylline in preventing leukocyte sequestration and lung injury caused by protamine at the end of cardiopulmonary bypass surgery.

BACKGROUND: Protamine has adverse effects on pulmonary gas exchange during the postoperative period. The objective of this study was to investigate the importance of aprotinin and pentoxifylline in preventing the leukocyte sequestration and lung injury caused by protamine administered after the termination of cardiopulmonary bypass (CPB). METHODS: Participants (n = 39) were allocated into three groups at the termination of CPB: Group 1, (control group, n = 16); Group 2 (aprotinin group, n = 12), who received protamine + aprotinin (15,000 IU/kg); and Group 3 (Pentoxifylline group, n = 11), who received protamine + pentoxifylline (10 mg/kg). Leukocyte counts in pulmonary and radial arteries were determined after the termination of CPB and before any drug was given (t1), and 5 minutes (t2), 2 hours (t3), 6 hours (t4) and 12 hours (t5) after the administration of protamine. Alveolar-arterial O2 gradient (A-aO2) and dynamic pulmonary compliance were measured at t1, t2 and t3. RESULTS: In the control group, an increase in pulmonary leukocyte sequestration was observed 5 minutes and 2 hours after protamine administration, after which this difference disappeared. No significant degree of pulmonary sequestration was detected in any measurements after protamine was administered in the aprotinin and pentoxifylline (PTX) groups. Dynamic lung compliance was 50.1, 45.2 and 47.2 ml/cm H2O in the control group, 49.2, 61.1 and 56.3 ml/cm H2O in the aprotinin group, and 49.5, 54.5 and 50.4 ml/cm H2O in the PTX group. The A-aO2 gradient was 212.2, 263.3 and 254.3 mm Hg in the control group, 209.4, 257.1 and 217.3 mm Hg in the aprotinin group, and 211.3, 260.8 and 219.2 mm Hg in the PTX group. CONCLUSION: Aprotinin and PTX treatments have favourable effects on lung function by reducing protamine-induced leukocyte sequestration into lungs at the end of CPB.