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Poult Sci 2008. 87:116-124. doi:10.3382/ps.2007-00160
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PHYSIOLOGY, ENDOCRINOLOGY, AND REPRODUCTION

Analysis of Plasma Serotonin Levels and Hemodynamic Responses Following Chronic Serotonin Infusion in Broilers Challenged with Bacterial Lipopolysaccharide and Microparticles1

M. E. Chapman*,2, R. L. Taylor{dagger} and R F. Wideman, Jr.*

* Department of Poultry Science, University of Arkansas, Fayetteville 72701; and {dagger} Department of Animal and Nutritional Sciences, University of New Hampshire, Durham 03824

2 Corresponding author: mchapman{at}uark.edu


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
There has been extensive interest in the role of serotonin (5-hydoxytryptamine, 5-HT) in the pathogenesis of pulmonary hypertension because episodes of pulmonary arterial hypertension in humans have been linked to serotoninergic appetite-suppressant drugs. In this study, we investigated the role of serotonin in the development of pulmonary hypertension induced by intravenously injecting bacterial lipopolysaccharide (LPS, endotoxin) and cellulose microparticles. In experiment 1, we used a 5-HT ELISA kit for the in vitro quantitative determination of 5-HT in plasma during the development of pulmonary hypertension induced by injecting LPS and cellulose microparticles i.v. in broilers. In experiment 2, broilers were either chronically infused with 5-HT via surgically implanted osmotic pumps or received sham surgery as a control. After a period of 10 d, the pulmonary arterial pressure was recorded during challenge with injected LPS or microparticles. Microparticles elicited 5-HT plasma levels more than 2-fold greater than those elicited by LPS from 15 to 45 min postinjection. This indicates that 5-HT is an important mediator in the pulmonary hypertensive response of broilers to microparticles, but may not play a prominent role in the pulmonary hypertensive response to LPS. Furthermore, chronic 5-HT infusion via osmotic pumps caused an increase in the duration of the pulmonary hypertensive response of broilers to microparticles, indicating that the infused 5-HT was sequestered by circulating thrombocytes and then released upon microparticle-mediated thrombocyte activation. Serotonin appears to play a less prominent role in the pulmonary hypertensive response of broilers to LPS, indicating that other mediators within the innate response to inflammatory stimuli may also be involved. These results are consistent with our hypothesis that pulmonary arterial hypertension ensues when vasoconstrictors such as 5-HT overwhelm the dilatory affects of vasodilators such as nitric oxide, thereby effectively reducing the pulmonary vascular capacity of pulmonary arterial hypertension-susceptible broilers.

Key Words: hypertension • broiler • serotonin • lipopolysaccharide • microparticle


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Fast-growing broilers are susceptible to pulmonary arterial hypertension (PAH), leading to pulmonary hypertension syndrome (ascites) when their right ventricle must progressively elevate the pulmonary arterial pressure (PAP) to propel the requisite cardiac output through lungs having a marginally inadequate pulmonary vascular capacity (Peacock et al., 1989; Julian, 1993; Wideman and Bottje, 1993; Wideman, 2000, Wideman et al., 2001). However, we still do not know what initiates PAH. Vaso-constriction, remodeling of the pulmonary vessel wall, and thrombosis are all thought to contribute to increased pulmonary vascular resistance in PAH. Any factor that increases the cardiac output, reduces the pulmonary vascular capacity, or triggers pulmonary vasoconstriction can contribute to the pathogenesis of PAH in broilers (Wideman, 2000), particularly when occurring within a genetic predisposition. Indeed, several cell types, including endothelial and smooth muscle cells, as well as inflammatory cells and thrombocytes, may play a significant role in PAH. There has been extensive interest in the role of serotonin (5-hydoxytrytamine, 5-HT) in the pathogenesis of pulmonary hypertension, because episodes of PAH in humans have been linked to serotoninergic appetite-suppressant drugs (Abenhaim et al., 1996). Increased plasma 5-HT levels have also been recorded in human patients who spontaneously developed idiopathic PAH (Hervé et al., 1995).

Serotonin is a potent pulmonary vasoconstrictor (Boe et al., 1980) and smooth muscle mitogen (Nemecek et al., 1986; Fanburg and Lee, 1997) that is synthesized from the essential amino acid Trp. It is actively accumulated by mammalian platelets and avian thrombocytes, where 98% or more of all circulating 5-HT is stored, and is released into the plasma during platelet or thrombocyte aggregation (Meyer and Sturkie, 1974; Cox, 1985; Lacoste-Eleaume et al., 1994). Serotonin is taken up by the 5-HT transporter (5-HTT) in several cells, including platelets and thrombocytes, as well as in pulmonary artery smooth muscle and endothelial cells. Serotonin has been implicated in the mechanisms responsible for pulmonary hypertension in several human, animal, and broiler studies (Douglas et al., 1981; Abenhaim et al., 1996; Chapman and Wideman, 2002). When platelets and thrombocytes aggregate, they release several physiologically active substances, including 5-HT, which causes proliferation of pulmonary vascular smooth muscle cells and stimulates vasoconstriction, thereby reducing the blood flow at the site of injury (Lee et al., 1994; Fanburg and Lee, 1997). Pulmonary vasoconstriction induced by 5-HT is believed to be mediated through 5-HT1B/1D and 5-HT2A receptors expressed by pulmonary smooth muscle cells, whereas the vascular remodeling associated with pulmonary hypertension appears to require the serotonin transporter (5-HTT). Therefore, determination of altered 5-HT plasma concentrations has great clinical significance for the assessment of broiler PAH.

In experiment 1, we used an enzyme immunoassay for the in vitro quantitative determination of plasma 5-HT during the development of pulmonary hypertension over a 12-h period, induced by injecting bacterial lipopolysaccharide (LPS) or cellulose microparticles i.v. in broilers. Lipopolysaccharide and microparticles cause pulmonary vasoconstriction and pulmonary hypertension in broilers (Wideman et al., 2001; Wideman and Erf, 2002), but the specific mediator of vasoconstriction has not been determined. Bacterial LPS can produce direct injury to the pulmonary vascular endothelium (Brigham and Meyrick, 1986; Meyrick et al., 1986); however, the pulmonary hypertension in gram-negative sepsis primarily results from LPS-activated host inflammatory responses (Brigham and Meyrick, 1986; Ghosh et al., 1993). Entrapped microparticles, in addition to physically occluding pulmonary arterioles, stimulate local tissues and leukocytes to release vasoactive substances capable of altering pulmonary vascular resistance by dilating or constricting the nearby vasculature (Wang et al., 2003; Wideman et al., 2004). Therefore, the objective of experiment 1 was to determine whether LPS- or microparticle-induced pulmonary hypertensive responses in broilers corresponded to increased plasma 5-HT levels. In experiment 2, broilers were either chronically infused with 5-HT via surgically implanted osmotic pumps or received sham surgery as a control (McCorkle and Taylor, 1989; Eddahibi et al., 1997). After a period of 10 d, the broilers were injected with LPS or microparticles while recording PAP. Experiment 2 had the objective of determining whether supplemental 5-HT infusions would exacerbate the pulmonary hypertensive response of broilers to LPS or microparticles.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Male broilers from a commercial hatchery (Cobb-Vantress Inc., Fayetteville, AR) were wing-banded and transported on the day of hatch (d 1) to the Poultry Environmental Research Laboratory at the University of Arkansas. They were placed on fresh wood shavings in environmental chambers (8 m2 of floor space) and were brooded at 33°C from d 1 to 5, 29°C from d 6 to 10, and 27°C from d 11 to 17. Thereafter, the broilers were maintained at 23.8°C until the experiment was terminated. The photoperiod was 24 h of light from d 1 to 5, and 23L:1D thereafter. Water was provided ad libitum via nipple type waterers. A corn-soybean meal starter ration (22.7% CP, 3,059 kcal of ME/kg, 1.5% Arg, and 1.43% Lys) was provided ad libitum and was formulated to meet minimum NRC (1994) standards for all ingredients. The diet was provided as crumbles during wk 1 and 2 and as pellets thereafter.

Experiment 1
From d 31 to 36, unanesthetized male broilers (n = 240; 1,904 ± 15.6 g of BW, mean ± SEM) were injected via the basilica vein with either 1 mg of Salmonella enterica serovar Typhimurium LPS (Sigma Chemical Co., St. Louis, MO; 0.5 mL dissolved at 2 mg/mL in a 0.9% sodium chloride solution, n = 120) or 0.35 mL of microgranular CM-32 ion-exchange cellulose microparticles (Fisher Scientific, St. Louis, MO), suspended at 0.02 g/mL in heparinized saline (n = 120), by using a 1-mL tuberculin syringe with a 22-gauge needle. Previous studies demonstrated that the l-mg dose of LPS elicits a maximal pulmonary hypertensive response in broilers (Wideman et al., 2001) without triggering endotoxin shock (Xie et al., 2000) and that approximately 0.3 to 0.35 mL of the 0.02 g/mL microparticle suspension is required to elevate the PAP of 6- to 7-wk-old male broilers significantly while triggering <10% mortality within 24 h postinjection (Wideman and Erf, 2002; Wideman et al., 2005b). At 15, 30, and 45 min, and 1, 2, 3, 4, 5, 6, 8, 10, and 12 h postinjection, 1.5-mL blood samples were collected from the basilica vein by using syringes containing 0.1 mL of a 0.9% sodium chloride solution containing 50 mg/mL of EDTA. The EDTA was used to chelate calcium and thereby prevent platelet activation and degranulation (Cox, 1985; Gobbi et al., 2003). The plasma was separated by centrifugation and was stored frozen at –4°C until analysis. Plasma 5-HT levels were measured by using a commercial enzyme immunoassay kit (Alpco Diagnostics, Windham, NH).

Experiment 2
Beginning on d 34 and continuing at daily intervals thereafter during the ensuing week, unanesthetized male broilers (n = 40; 2,307 ± 139.1 g of BW, mean ± SEM) were placed on a surgical board and restrained in dorsal recumbency. The feathers were removed from the dorsal surface of the neck as needed, intracutaneous injections of 2% lidocaine HCl were administered as a local anesthetic, and an incision was made adjacent to the site chosen for the subcutaneous surgical implantations. Osmotic pumps (2ML2 Alzet, Durect Corporation, Cupertino, CA) containing 2 mL of 5-HT (Sigma Chemical Co.) at 10 mg/mL in 0.9% saline (pump group, n = 20) or gelatin capsules (sham group, n = 20) were implanted and the implantation site was then closed with wound clips. The 2ML2 Alzet osmotic pumps are designed to deliver a test drug at the consistent rate of 5 µL/h for 14 d. Based on a previous study (Chapman and Wideman, 2002) the 10 mg/mL concentration of 5-HT was chosen to increase 5-HT levels in circulating thrombocytes without causing the pulmonary vasoconstriction leading to PAH.

Ten to 12 d postimplantation (starting on d 44), the broilers were anesthetized to a light surgical plane with intramuscular injections of allobarbitol (5,5-diallylbarbituric acid, 3.0 mL, 25 mg/mL, Sigma Chemical Co.) and ketamine HCl (1.0 mL, 100 mg/mL, Bedford Laboratories, Bedford, OH). The birds were placed on a heated surgical board (30°C) and restrained in dorsal recumbency. The left wing was extended and feathers were removed from the ventral surface as needed to uncover the skin over the basilica vein. After intracutaneous injections of 2% lidocaine HCl were administered as a local anesthetic, an incision was made to expose the vein, which then was cannulated with a Silastic catheter (0.03 cm i.d., 0.09 cm o.d.) filled with a 0.9% sodium chloride solution containing 200 IU of heparin/mL. The catheter was attached to a blood pressure transducer interfaced through a Transbridge preamplifier (World Precision Instruments, Sarasota, FL) to a Biopac MP 100 data acquisition system using AcqKnowledge software (Biopac Systems Inc., Goleta, CA). The catheter was advanced through the right atrium and ventricle into a pulmonary artery while the characteristic pulse pressures were monitored to identify the location (Wideman et al., 1996; Chapman and Wideman, 2001). All pulmonary arterial pressure readings were made with the transducer at the level of the thoracic inlet. Prior to recording, the system was calibrated in millimeters of mercury (mmHg) by using a mercury manometer. The left basilica vein was also cannulated with PE-50 polyethylene tubing filled with heparinized saline for i.v. injections. Control data were recorded for 10 min after surgical preparations were complete and a stabilization period of 10 min had elapsed. The osmotic pump and sham groups were then injected i.v. with 1 mg of Salmonella Typhimurium LPS (0.5 mL dissolved at 2 mg/mL in a 0.9% sodium chloride solution, n = 20) or 0.35 mL of cellulose microparticles suspended at 0.02 g/mL in heparinized saline (n = 20). Data were recorded for a further 60 min, after which the birds were euthanized with 10 mL i.v. of 0.1 M KCl.

Data Analysis
The primary channel of the Biopac MP 100 data acquisition system recorded PAP (mmHg). These data was averaged electronically during representative sample intervals while accommodating for the influences of pulse pressure and respiratory cycles on pulmonary arterial pressure (Wideman et al., 1996). Data were analyzed by t-test (comparison between 2 groups within a single sample interval) or within a group over time (across sample intervals) by using the SigmaStat Repeated Measures ANOVA procedure (Jandel Scientific, 1994). The Dunnett method was used for the separation of treatment means. The threshold for significance in all cases was P ≤ 0.05.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Experiment 1
At 15, 30, and 45 min postinjection, microparticles elicited plasma levels of 5-HT of 171 ± 40, 234 ± 80, and 175 ± 50 ng/mL, respectively during the pulmonary arterial hypertensive response in broilers (Figure 1Go). This 5-HT response to microparticles was more than 2-fold greater (P = 0.04) than the responses elicited by LPS (69 ± 20, 31 ± 13, and 80 ± 50 ng/mL, respectively). With the exception of a significant difference at the 6-h sample interval, the plasma 5-HT levels of broilers injected with microparticles were similar to 5-HT levels observed in broilers injected with LPS. The 5-HT concentrations in microparticle-injected birds declined after the 30-min measurement.


Figure 1
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  		Figure 1. Plasma serotonin levels (ng/mL) for male broilers (n = 240; 1,904 ± 15.6 g of BW, mean ± SEM) in experiment 1, at 15, 30, and 45 min and at 1, 2, 3, 4, 5, 6, 8, 10, and 12 h after lipopolysaccharide (LPS) or microparticle injection. Subscript letters (a and b) represent differences (P ≤ 0.05) between groups within individual sample intervals. Asterisks (*) represent differences (P ≤ 0.05) within a group across sample intervals. 5-HT = 5-hydroxytryptamine (serotonin).

 
Experiment 2
For broilers that subsequently were injected with LPS, the initial baseline PAP averaged approximately 20 mmHg in the broilers from the sham group, whereas after 10 d of chronic 5-HT infusion via osmotic pumps, the baseline PAP averaged approximately 23 mmHg in the pump group (sample intervals St to S10; Figure 2aGo). The numerically greater initial baseline PAP values for broilers in the pump group apparently shifted the group’s absolute PAP response to LPS upward by approximately 3 mmHg when compared with the absolute PAP response to LPS by the sham group (sample intervals T to T60). Accordingly, absolute PAP values for both groups were recalculated to reflect the LPS-induced percentage change from the initial baseline PAP values, as shown in Figure 2bGo. Injecting 1 mg of LPS into the wing vein caused the PAP to rise within 10 min and reach a peak of approximately 70% above baseline values within 25 min (P > 0.05) in both groups of broilers, regardless of treatment during the preceding 10 d (5-HT infusion, sham controls). The percentage change in PAP then subsided toward baseline values by the end of the experiment in both the 5-HT-treated and control groups. At no sample interval during the hypertensive response to LPS (T to T60) did the percentage change in PAP differ between the groups (P > 0.05).


Figure 2
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  		Figure 2. a) Pulmonary arterial pressure (PAP) and b) percentage change from the initial PAP (PAP % change) for male broilers (n = 40; 2,307 ± 139.1 g of BW, mean ± SEM) in experiment 2, at the start of data collection and at intervals of 5 min thereafter (St, S5, S10), within 30 s after lipopolysaccharide (LPS) injection and at intervals of 5 min thereafter (T to T60). Asterisks (*) represent differences (P ≤ 0.05) within a group across sample intervals.

 
For broilers that were subsequently injected with microparticles, the initial baseline PAP was approximately 16.5 mmHg in birds from the sham group and approximately 18.5 mmHg in birds that had received chronic 5-HT infusion via osmotic pumps (sample intervals St to S10; Figure 3aGo). Because of the numerically greater initial absolute PAP values in broilers from the pump group when compared with the sham broilers, the percentage change from the initial PAP was calculated as shown in Figure 3bGo for comparisons of the pulmonary arterial hypertensive response between the groups. Injecting microparticles into the wing vein caused the PAP to rise within 5 min to a peak of approximately 50% above baseline values (P > 0.05) in broilers, regardless of treatment (sample interval TPk, sham and pump groups). The percentage change in PAP remained elevated by approximately 20% above baseline values by the end of the experiment (sample interval T60) in broilers that were chronically infused with 5-HT, whereas in broilers from the sham group, the pulmonary hypertensive response subsided 20 min earlier (sample interval T40) to a level that did not differ from the initial baseline values. At no sample interval during the hypertensive response to micro-particles (T to T60) did the absolute PAP or percentage change in PAP differ between the groups (P > 0.05).


Figure 3
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  		Figure 3. a) Pulmonary arterial pressure (PAP) and b) percentage change from the initial PAP (PAP % change) for male broilers (n = 40; 2,307 ± 139.1 g of BW, mean ± SEM) in experiment 2, at the start of data collection and at intervals of 5 min thereafter (St, S5, S10), within 30 s after microparticle injection and at intervals of 5 min thereafter (T to T60). Asterisks (*) represent differences (P ≤ 0.05) within a group across sample intervals.

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
Previously, we demonstrated that the nonselective 5-HT1/2 receptor antagonist methiothepin virtually eliminated the pulmonary hypertensive response of broilers to i.v. microparticle injections but had minimal impact on LPS-mediated pulmonary hypertension (Chapman and Wideman, 2006b). However, changes in plasma 5-HT levels following LPS or microparticle injections had not previously been demonstrated. In experiment 1 at 15 to 45 min postinjection, the microparticles elicited plasma levels of 5-HT more than 2-fold greater than those elicited by LPS in broilers. Plasma 5-HT levels then declined over the remainder of the experiment toward levels similar to those observed in broilers injected with LPS. The present and previous studies have consistently indicated that 5-HT is an important mediator of broilers’ pulmonary hypertensive response to microparticles; however, 5-HT may not play a prominent role in the pulmonary hypertensive response of broilers to LPS. This suggests that other mediators of the innate immune response interact with LPS.

It has been shown that within minutes after being injected, entrapped microparticles are surrounded by focal aggregates of thrombocytes and monocytes or macrophages (Wideman et al., 2002; Wang et al., 2003), which potentially can trigger these leukocytes and the adjacent vascular endothelium to synthesize and release potent vasoactive compounds within close proximity to the vascular smooth muscle (Wideman, 2001; Wideman et al., 2004). The tendency for elevated plasma 5-HT levels to wane within 1 h postmicroparticle injection, despite the persistence of cellulose microparticles within the pulmonary microvasculature for more than 10 d (Wideman et al., 2002; Wang et al., 2003), may be attributable to exhaustion of thrombocyte 5-HT reserves, cessation of thrombocyte activation, or inhibition by nitric oxide (NO; Moro et al., 1995). The available 5-HT also may be internalized into pulmonary endothelial cells via the 5-HTT. Pulmonary arterial hypertension is characterized by vasoconstriction, followed by structural remodeling associated with smooth muscle cell proliferation, both of which can be caused by increases in plasma 5-HT. Pulmonary vasoconstriction is thought to be mediated by 5-HT2A and 5-HT1B/1D receptors expressed on vascular smooth muscle cells (Choi and Maroteaux, 1996; MacLean et al., 1996; Keegan et al., 2001). Structural remodeling is mediated by 5-HTT expressed on vascular smooth muscle cells in the lungs. Serotonin translocated into the smooth muscle cells by 5-HTT is mitogenic, causing hypertrophy and proliferation of the medial muscle layer in pulmonary arterioles (Lee et al., 1991, 1994; Ramamoorthy et al., 1993; Fanburg and Lee, 1997; Eddahibi et al., 1999).

Serotonin, it seems, may not play a prominent role in the pulmonary hypertensive response to LPS in broilers, suggesting that the pathological progression toward PAH has more than one progenitor. The vasoconstrictor thromboxane A2 (TxA2) has been shown to increase pulmonary vascular resistance and PAP in broilers (Wideman et al., 1998a, 1999a, 2001), and several studies have associated TxA2 with the pulmonary hypertension during endotoxemia. Levels of thromboxane B2 (a stable metabolite of TxA2) in plasma and lung lymph increased acutely after LPS administration in several mammalian species (Ball et al., 1983; Snapper et al., 1983; Kubo and Kobayashi, 1985). Furthermore, the fawn-hooded rat, which spontaneously develops PAH, is believed to be a model of 5-HT-mediated PAH owing to a platelet storage defect that impairs uptake and retention of serotonin and elevates plasma serotonin levels (Tschopp and Zucker, 1972; Gonzalez et al., 1998). However, recent studies have implicated overexpression of endothelin-1 (ET-1) mRNA and overproduction of the pulmonary vasoconstrictor ET-1 peptide in fawn-hooded rat lung tissues as the potential mediator of PAH (Hahn et al., 1990; Nagaoka et al., 2001). Additional studies using fawn-hooded rat lungs point toward impaired endothelial NO synthase expression (Tyler et al., 1999), leading to sustained pulmonary vaso-constriction and PAH. Endothelial dysfunction has been identified as playing a key role in PAH pathophysiology, leading to impaired release of NO and prostacyclin and an overexpression of vasoconstrictors such as thromboxane and endothelin-1 (Humbert et al., 2004). Pulmonary hypertension therefore appears to have a multifactorial pathology.

In experiment 2, chronic (10-d) 5-HT infusion via osmotic pumps caused a numerical increase in PAP in comparison with control broilers that had received sham surgery, suggesting a role for 5-HT in maintaining the basal tone of the pulmonary arterial wall in broilers. These results are consistent with the observation that the 5-HT receptor blocker methiothepin causes a 25% reduction in baseline PAP in broilers (Chapman and Wideman, 2006a,b). The peak PAP response to 1 mg of LPS attained a similar magnitude of 70% above baseline values within 25 min postinjection in both 5-HT-infused and sham groups, providing further evidence that 5-HT may not play a prominent role in the pulmonary hypertensive response to LPS in broilers. Injecting microparticles into the wing vein caused the PAP to rise within 5 min to a peak of approximately 50% above baseline values in both 5-HT-infused and control broilers. Thereafter, the PAP remained elevated by approximately 20% above baseline values for the duration of the experiment in broilers chronically infused with 5-HT but not in broilers from the sham group. These observations are consistent with the hypothesis that the infused 5-HT was sequestered by circulating thrombocytes via the 5-HTT and then released upon microparticle-mediated thrombocyte activation. Endothelial NO synthase can be induced in endothelial cells to produce the vasodilator NO in response to increased rates in blood flow through constricted vascular channels. Previous studies have demonstrated that NO produced by endothelial NO synthase likely contributes to the attenuation of the pulmonary hypertensive responses of broilers to LPS and microparticles seen toward the end of experiment 2 (Bowen et al., 2006; Wideman et al., 2006).

In summary, this study provides direct evidence that 5-HT plays an important role in the pulmonary hypertensive response to intravenously injected microparticles in broilers. Serotonin appears to play a less prominent role in the pulmonary hypertensive response of broilers to intravenously injected LPS, indicating that other mediators within the innate response to inflammatory stimuli, such as TxA2, ET-1, or NO, may also be involved. The current observations support our hypothesis that pulmonary hypertension syndrome ensues when vasoconstrictors overwhelm the dilatory affects of vasodilators such as NO.


   ACKNOWLEDGMENTS
 
This project was supported by the National Research Initiative of the USDA Cooperative State Research, Education and Extension Service (Washington, DC), USDA/CSREES/NRI grant no. 2003-35204-13392, and by an Animal Health Grant from the University of Arkansas Agricultural Experiment Station (Fayetteville, AR).


   FOOTNOTES
 
1 United States patent no. 6,720,473 protects the exclusive rights of the University of Arkansas to all uses of the intravenous micro-particle injection technology within the context of evaluating or affecting pulmonary vascular capacity, pulmonary vascular resistance, pulmonary hypertension, cardio-pulmonary hemodynamics, and susceptibility to pulmonary hypertension and pulmonary hypertension syndrome (ascites) in domesticated animal species. Back

Received for publication April 18, 2007. Accepted for publication September 10, 2007.


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 ABSTRACT
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 RESULTS
 DISCUSSION
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R. F. Wideman, O. T. Bowen, and G. F. Erf
Broiler pulmonary hypertensive responses during lipopolysaccharide-induced tolerance and cyclooxygenase inhibition
Poult. Sci., January 1, 2009; 88(1): 72 - 85.
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