We evaluated the pharmacokinetics of IV administered sodium heparin and the pharmacodynamic effect of heparin on lipoprotein lipase (LPL) activity Horses were allotted to 3 groups. Plasma samples were obtained from each horse before and at various times for 6 hours after heparin administration for determination of heparin concentration, LPL activity, and activated partial thromboplastin time (APTT). The disposition of heparin was dose dependent. The area under the plasma heparin concentration vs time curve (AUC) increased more than proportionally with dose, indicating that heparin elimination was nonlinear. Total clearance of heparin was similar after the 40 and 80 IU/kg of body weight dosages, averaging 0.45 and 0.36 IU/kg/min, respectively. However, after administration of the 120 IU/kg dose, clearance was significantly less than that after the 40 IU/kg dose. The half-life of heparin averaged 53, 70, and 136 minutes after 40, 80, and 120 IU/kg, respectively, with significant differences observed between the low and high doses. In contrast to heparin, the area under the plasma concentration vs time curve for LPL activity increased less than proportionally with dose. Maximal LPL activity observed was independent of dose, averaging 4.8 micromole of free fatty acids/ml/h. The APTT was significantly prolonged for 120 minutes after administration of the 40 IU/kg dose. Correlation coefficients for LPL activity vs either plasma heparin concentration or APTT were less than 0.7, indicating that neither laboratory measure can be used to accurately predict plasma LPL activity.

We evaluated the pharmacokinetics of IV administered sodium heparin and the pharmacodynamic effect of heparin on lipoprotein lipase (LPL) activity Horses were allotted to 3 groups. Plasma samples were obtained from each horse before and at various times for 6 hours after heparin administration for determination of heparin concentration, LPL activity, and activated partial thromboplastin time (APTT). The disposition of heparin was dose dependent. The area under the plasma heparin concentration vs time curve (AUC) increased more than proportionally with dose, indicating that heparin elimination was nonlinear. Total clearance of heparin was similar after the 40 and 80 IU/kg of body weight dosages, averaging 0.45 and 0.36 IU/kg/min, respectively. However, after administration of the 120 IU/kg dose, clearance was significantly less than that after the 40 IU/kg dose. The half-life of heparin averaged 53, 70, and 136 minutes after 40, 80, and 120 IU/kg, respectively, with significant differences observed between the low and high doses. In contrast to heparin, the area under the plasma concentration vs time curve for LPL activity increased less than proportionally with dose. Maximal LPL activity observed was independent of dose, averaging 4.8 micromole of free fatty acids/ml/h. The APTT was significantly prolonged for 120 minutes after administration of the 40 IU/kg dose. Correlation coefficients for LPL activity vs either plasma heparin concentration or APTT were less than 0.7, indicating that neither laboratory measure can be used to accurately predict plasma LPL activity.

We compared the ability of 2 alpha2-adrenergic receptor antagonists, atipamezole and yohimbine, to reverse medetomidine-induced CNS depression and cardiorespiratory changes in lambs. Twenty lambs (7.8 +/- 2.6 kg) were randomly allotted to 4 treatment groups (n = 5). Each lamb was given medetomidine (30 micrograms/kg of body weight, IV), followed in 15 minutes by IV administration of atipamezole (30 or 60 micrograms/kg), yohimbine (1 mg/kg), or 0.9% NaCl (saline) solution. Medetomidine caused lateral recumbency in 1 to 2 minutes in all treated lambs. Medetomidine significantly (P < 0.05) decreased heart rate at 5 and 10 minutes after its administration. Heart rate remained above 120 beats/min, and severe bradycardia (< 70 beats/min) and other arrhythmias did not occur throughout the study. Medetomidine also induced tachypnea in all treated lambs. The tachypnea was abolished by atipamezole and yohimbine, but not by saline solution administration. The medetomidine-induced tachypnea did not significantly affect arterial pH and PaCO2. Arterial oxygen tension was within acceptable range (PaO2 = 71 to 62 mm of Hg), but was lower than expected. Administration of atipamezole, yohimbine, or saline solution did not change PaO2 significantly. Lambs treated with 30 or 60 micrograms of atipamezole/kg were able to walk unassisted in 2.4 +/- 0.4 and 2.3 +/- 0.7 minutes, respectively, whereas yohimbine-and saline-treated lambs did not walk unassisted until 15.6 +/- 2.7 and 73.0 +/- 6.8 minutes later, respectively. Results of this study indicated that medetomidine is a potent CNS depressant in lambs. Atipamezole at dosage of 30 or 60 micrograms/kg was equally effective, and was more effective in antagonizing medetomidine-induced CNS depression than was yohimbine.

We compared the ability of 2 alpha2-adrenergic receptor antagonists, atipamezole and yohimbine, to reverse medetomidine-induced CNS depression and cardiorespiratory changes in lambs. Twenty lambs (7.8 +/- 2.6 kg) were randomly allotted to 4 treatment groups (n = 5). Each lamb was given medetomidine (30 micrograms/kg of body weight, IV), followed in 15 minutes by IV administration of atipamezole (30 or 60 micrograms/kg), yohimbine (1 mg/kg), or 0.9% NaCl (saline) solution. Medetomidine caused lateral recumbency in 1 to 2 minutes in all treated lambs. Medetomidine significantly (P < 0.05) decreased heart rate at 5 and 10 minutes after its administration. Heart rate remained above 120 beats/min, and severe bradycardia (< 70 beats/min) and other arrhythmias did not occur throughout the study. Medetomidine also induced tachypnea in all treated lambs. The tachypnea was abolished by atipamezole and yohimbine, but not by saline solution administration. The medetomidine-induced tachypnea did not significantly affect arterial pH and PaCO2. Arterial oxygen tension was within acceptable range (PaO2 = 71 to 62 mm of Hg), but was lower than expected. Administration of atipamezole, yohimbine, or saline solution did not change PaO2 significantly. Lambs treated with 30 or 60 micrograms of atipamezole/kg were able to walk unassisted in 2.4 +/- 0.4 and 2.3 +/- 0.7 minutes, respectively, whereas yohimbine- and saline-treated lambs did not walk unassisted until 15.6 +/- 2.7 and 73.0 +/- 6.8 minutes later, respectively. Results of this study indicated that medetomidine is a potent CNS depressant in lambs. Atipamezole at dosage of 30 or 60 micrograms/kg was equally effective, and was more effective in antagonizing medetomidine-induced CNS depression than was yohimbine.

Cytologic examination of bronchoalveolar lavage fluid (BALF), including phenotypic analysis of lymphocytes, was performed on 32 Standardbreds with poor race performance and endoscopic examination findings characteristic of inflammatory airway disease (IAD). Nucleated cell counts in BALF from IAD-affected horses were higher than those in control horses; the cytologic profile of BALF in affected horses included mixed inflammation, characterized by mild neutrophilia, lymphocytosis, and monocytosis. Eosinophil and mast cell counts were not higher in the IAD-affected group, compared with those in the control group; however, 4 IAD-affected horses had marked eosinophilia (24.7 +/- 4.8% SEM) in BALF. Phenotypic analysis of lymphocytes in BALF obtained from IAD-affected horses revealed a low proportion of CD4-positive cells and B cells, compared with those in the control group; these findings may have been representative of a greater proportion of non-B, non-T cells (null cells) in horses with IAD. The cytologic profile of BALF obtained from horses with IAD differed from that in horses affected with chronic obstructive pulmonary disease, suggesting that the pathogenesis of inflammation in horses with IAD may differ from that of chronic obstructive pulmonary disease.

Cytologic examination of bronchoalveolar lavage fluid (BALF), including phenotypic analysis of lymphocytes, was performed on 32 Standardbreds with poor race performance and endoscopic examination findings characteristic of inflammatory airway disease (IAD). Nucleated cell counts in BALF from IAD-affected horses were higher than those in control horses; the cytologic profile of BALF in affected horses included mixed inflammation, characterized by mild neutrophilia, lymphocytosis, and monocytosis. Eosinophil and mast cell counts were not higher in the IAD-affected group, compared with those in the control group; however, 4 IAD-affected horses had marked eosinophilia (24.7 +/- 4.8% SEM) in BALF. Phenotypic analysis of lymphocytes in BALF obtained from IAD-affected horses revealed a low proportion of CD4-positive cells and B cells, compared with those in the control group; these findings may have been representative of a greater proportion of non-B, non-T cells (null cells) in horses with IAD. The cytologic profile of BALF obtained from horses with IAD differed from that in horses affected with chronic obstructive pulmonary disease, suggesting that the pathogenesis of inflammation in horses with IAD may differ from that of chronic obstructive pulmonary disease.

Adrenocortical function was assessed in 27 Beagle pups at 2, 4, 6, 8, 10, and 12 weeks of age by determination of plasma sodium, potassium, and chloride concentrations; serum aldosterone and cortisol concentrations; and plasma ACTH concentrations. Serum cortisol concentration was measured before and 1 and 2 hours after IM administration of 2.2 IU of ACTH/kg of body weight. Serum progesterone concentration also was determined for all pups at 2, 4, and 6 weeks of age. Mean baseline cortisol concentration was lower for pups 8 weeks old or younger than for mature dogs. Nevertheless, mean serum ACTH-stimulated cortisol concentration in dogs of all age groups increased into the adult reference range after administration of ACTH. For pups 4 weeks old or younger, increase in cortisol concentration was maximal at 2 hours after ACTH administration. However, in pups between 6 and 12 weeks of age, the increase in cortisol concentration was maximal 1 hour after ACTH administration in about a third of the pups, whereas the remaining pups had peak values at 2 hours. Mean plasma sodium, potassium, and chloride concentrations for each age group were within the reference ranges established for mature dogs, with the exception of lower mean plasma sodium and chloride concentrations in pups 4 weeks old or younger. Mean serum aldosterone concentration in pups of each age group was substantially higher than the range of aldosterone concentrations for clinically normal mature dogs. Median progesterone concentration was uniformly less than 0.2 ng/ml for all pups 6 weeks old or younger. The normal endogenous ACTH concentration and adequate cortisol responses to exogenous ACTH seen in our pups would support functional pituitary gland and adrenal cortex for cortisol production. The low baseline cortisol concentration observed in the pups of this study may be related to reduced binding of cortisol to plasma proteins, as exists in human infants.

Adrenocortical function was assessed in 27 Beagle pups at 2, 4, 6, 8, 10, and 12 weeks of age by determination of plasma sodium, potassium, and chloride concentrations; serum aldosterone and cortisol concentrations; and plasma ACTH concentrations. Serum cortisol concentration was measured before and 1 and 2 hours after IM administration of 2.2 IU of ACTH/kg of body weight. Serum progesterone concentration also was determined for all pups at 2, 4, and 6 weeks of age. Mean baseline cortisol concentration was lower for pups 8 weeks old or younger than for mature dogs. Nevertheless, mean serum ACTH-stimulated cortisol concentration in dogs of all age groups increased into the adult reference range after administration of ACTH. For pups 4 weeks old or younger, increase in cortisol concentration was maximal at 2 hours after ACTH administration. However, in pups between 6 and 12 weeks of age, the increase in cortisol concentration was maximal 1 hour after ACTH administration in about a third of the pups, whereas the remaining pups had peak values at 2 hours. Mean plasma sodium, potassium, and chloride concentrations for each age group were within the reference ranges established for mature dogs, with the exception of lower mean plasma sodium and chloride concentrations in pups 4 weeks old or younger. Mean serum aldosterone concentration in pups of each age group was substantially higher than the range of aldosterone concentrations for clinically normal mature dogs. Median progesterone concentration was uniformly less than 0.2 ng/ml for all pups 6 weeks old or younger. The normal endogenous ACTH concentration and adequate cortisol responses to exogenous ACTH seen in our pups would support functional pituitary gland and adrenal cortex for cortisol production. The low baseline cortisol concentration observed in the pups of this study may be related to reduced binding of cortisol to plasma proteins, as exists in human infants. The hyponatremia and increased aldosterone concentration may be explained by reduced renal tubular response to aldosterone, as also evidenced in the human infant kidney.

Because caffeine is metabolized by the hepatic P-450 cytochrome oxidase system, clearance of caffeine is an excellent quantitative test of hepatic function in human beings. It is currently used in much the same way that creatinine clearance is used to assess renal function. Caffeine clearance was measured in lactating dairy cows initially to determine the suitability of caffeine clearance as an indicator of hepatic function in cattle. Pharmacokinetic variables of caffeine were studied in 6 adult lactating dairy cows after IV administration of a single dose of caffeine sodium benzoate (2 mg of caffeine/kg of body weight). Caffeine concentration was analyzed by use of an automated enzyme immunoassay. The lower limit of detection of the assay for caffeine in serum was 0.079 micrograms/ml. Serum caffeine concentration-time curves best fit an open two-compartment pharmacokinetic model. Harmonic mean elimination half-life was 3.8 (range, 2.6 to 6.9) hours, and total clearance was 0.118 (range, 0.090 to 0.197) L/kg/h. Milk caffeine concentration was similar to serum concentration 1.5 to 24 hours after caffeine administration. Adverse effects were not observed in cows given caffeine.

Because caffeine is metabolized by the hepatic P-450 cytochrome oxidase system, clearance of caffeine is an excellent quantitative test of hepatic function in human beings. It is currently used in much the same way that creatinine clearance is used to assess renal function. Caffeine clearance was measured in lactating dairy cows initially to determine the suitability of caffeine clearance as an indicator of hepatic function in cattle. Pharmacokinetic variables of caffeine were studied in 6 adult lactating dairy cows after IV administration of a single dose of caffeine sodium benzoate (2 mg of caffeine/kg of body weight). Caffeine concentration was analyzed by use of an automated enzyme immunoassay. The lower limit of detection of the assay for caffeine in serum was 0.079 micrograms/ml. Serum caffeine concentration-time curves best fit an open two-compartment pharmacokinetic model. Harmonic mean elimination half-life was 3.8 (range, 2.6 to 6.9) hours, and total clearance was 0.118 (range, 0.090 to 0.197) L/kg/h. Milk caffeine concentration was similar to serum concentration 1.5 to 24 hours after caffeine administration. Adverse effects were not observed in cows given caffeine.

Free, autogenous, full-thickness skin grafts were applied to 10 dogs; 5 dogs were given an iron chelator, deferoxamine-10% hydroxyethyl pentafraction starch (DEF-HES; 50 mg/kg of body weight, IV), and 5 dogs were given an equal volume of 10% hydroxyethyl pentafraction starch (HES) in 0.9% saline solution (5 ml/kg, IV). All dogs (DEF-HES/HBO- and HES/HBO-treated) were exposed to 60 minutes of hyperbaric oxygen (HBO) at 2 atmospheres absolute pressure twice daily for 10 days, beginning the day of surgery. The percentage of viable graft on day 10 was lower in HES/HBO-treated-dogs (mean +/- SD, 13.3 +/- 21.3%; median, 3.0%) than in DEF-HES/HBO-treated dogs (64.7 +/- 39.2%; 88.3%; P = 0.095, Mann-Whitney two-tailed test). There was a positive correlation between percentage of viable graft (on day 10) and percentage of haired skin on the graft site (on day 28) for all dogs (r = 0.91) and for HES/HBO-treated dogs (r = 0.97). The DEF-HES/HBO-treated dogs had less consistent correlation (r = 0.67). Perivascular aggregates of foamy cells were observed in the superficial and reticular portions of the dermis and in the subcutaneous tissue on both surfaces of the panniculus muscle in the graft sites of DEF-HES/HBO-treated dogs. These cells were also observed in the dermis, but not subcutaneous tissue of the control skin sections, and in some viscera of DEF-HES/HBO-treated dogs. Deferoxamine appears to attenuate the detrimental effect of HBO and HES on survival of free skin grafts. However, clinical use of HBO is not recommended as adjunct treatment for free skin grafts in dogs in the first 10 days after grafting. Administration of DEF-HES is not recommended because it has failed to improve the survival of free skin grafts, and the consequence of the cellular response seen in this study is undetermined.

Free, autogenous, full-thickness skin grafts were applied to 10 dogs; 5 dogs were given an iron chelator, deferoxamine-10% hydroxyethyl pentafraction starch (DEF-HES; 50 mg/kg of body weight, IV), and 5 dogs were given an equal volume of 10% hydroxyethyl pentafraction starch (HES) in 0.9% saline solution (5 ml/kg, IV). All dogs (DEF-HES/HBO- and HES/HBO-treated) were exposed to 60 minutes of hyperbaric oxygen (HBO) at 2 atmospheres absolute pressure twice daily for 10 days, beginning the day of surgery. The percentage of viable graft on day 10 was lower in HES/HBO-treated-dogs (mean +/- SD, 13.3 +/- 21.3%; median, 3.0%) than in DEF-HES/HBO-treated dogs (64.7 +/- 39.2%; 88.3%; P = 0.095, Mann-Whitney two-tailed test). There was a positive correlation between percentage of viable graft (on day 10) and percentage of haired skin on the graft site (on day 28) for all dogs (r = 0.91) and for HES/HBO-treated dogs (r = 0.97). The DEF-HES/HBO-treated dogs had less consistent correlation (r = 0.67). Perivascular aggregates of foamy cells were observed in the superficial and reticular portions of the dermis and in the subcutaneous tissue on both surfaces of the panniculus muscle in the graft sites of DEF-HES/HBO-treated dogs. These cells were also observed in the dermis, but not subcutaneous tissue of the control skin sections, and in some viscera of DEF-HES/HBO-treated dogs. Deferoxamine appears to attenuate the detrimental effect of HBO and HES on survival of free skin grafts. However, clinical use of HBO is not recommended as adjunct treatment for free skin grafts in dogs in the first 10 days after grafting. Administration of DEF-HES is not recommended because it has failed to improve the survival of free skin grafts, and the consequence of the cellular response seen in this study is undetermined.

Immunoglobulin E is produced in response to parasitic nematodes that undergo blood and tissue migrations. Results of our previous studies indicated that IgE and IgG respond to Dirofilaria immitis in experimentally infected dogs. To determine the association between treatment with the larvicide, diethylcarbamazine (DEC), and antibody responses and to examine the potential influence of infection with a nonfilarid intestinal nematode on isotype-specific immune responses, we monitored, by use of isotype-specific ELISA, separate IgE and IgG responses against D immitis in 4 groups (A-D) of 8 dogs experimentally coinfected with D immitis and Ancylostoma caninum. All dogs were monitored from 2 weeks before inoculation with D immitis, through postinoculation (PI) week 20. Group-B dogs received a daily regimen of 6.6 mg of DEC/kg of body weight. Group-C dogs received 4.95 mg of oxibendazole/kg daily. Group-D dogs received DEC and oxibendazole, equivalent to the daily doses given to dogs of groups B and C. All dogs given oxibendazole had no A caninum at necropsy. Of the groups receiving DEC, 3 group-B dogs each had 1 to 2 D immitis at necropsy. When results of chronologic IgE determination for all groups were statistically compared, only groups B and C had significant (P = 0.0148 and P << 0.00005, respectively) increases in IgE values. Group-C dogs had the highest IgE values from PI week 10 until the end of the study, whereas IgG values were statistically identical to those of group-A dogs. Group-B dogs given only DEC and having the least number of D immitis of all groups, had IgE values that peaked at PI week 6; values were significantly (P = 0.0002) higher than those for all other groups. In Group-B dogs, IgG values increased significantly (P << 0.00005) only at PI week 20 and were significantly (P << 0.00005) decreased after PI week 6, compared with values for all other groups. Group D containing 6 dogs infected with 1 to 18 D immitis found at necropsy had IgE values betwee.

Immunoglobulin E is produced in response to parasitic nematodes that undergo blood and tissue migrations. Results of our previous studies indicated that IgE and IgG respond to Dirofilaria immitis in experimentally infected dogs. To determine the association between treatment with the larvicide, diethylcarbamazine (DEC), and antibody responses and to examine the potential influence of infection with a nonfilarid intestinal nematode on isotype-specific immune responses, we monitored, by use of isotype-specific ELISA, separate IgE and IgG responses against D immitis in 4 groups (A-D) of 8 dogs experimentally coinfected with D immitis and Ancylostoma caninum. All dogs were monitored from 2 weeks before inoculation with D immitis, through postinoculation (PI) week 20. Group-B dogs received a daily regimen of 6.6 mg of DEC/kg of body weight. Group-C dogs received 4.95 mg of oxibendazole/kg daily. Group-D dogs received DEC and oxibendazole, equivalent to the daily doses given to dogs of groups B and C. All dogs given oxibendazole had no A caninum at necropsy. Of the groups receiving DEC, 3 group-B dogs each had 1 to 2 D immitis at necropsy. When results of chronologic IgE determination for all groups were statistically compared, only groups B and C had significant (P = 0.0148 and P << 0.00005, respectively) increases in IgE values. Group-C dogs had the highest IgE values from PI week 10 until the end of the study, whereas IgG values were statistically identical to those of group-A dogs. Group-B dogs given only DEC and having the least number of D immitis of all groups, had IgE values that peaked at PI week 6; values were significantly (P = 0.0002) higher than those for all other groups. In Group-B dogs, IgG values increased significantly (P << 0.00005) only at PI week 20 and were significantly (P << 0.00005) decreased after PI week 6, compared with values for all other groups. Group D containing 6 dogs infected with 1 to 18 D immitis found at necropsy had IgE values between those of group-B dogs and those of non-DEC-treated groups at PI week 6. There was no difference in IgG values between 3 groups at PI week 6, and IgE values were found to be a better correlator than were IgG values to the number of D immitis larvae killed in the tissues during this period. All differences in IgG and IgE values not only correlated with treatment status and number of D immitis adults found at necropsy, but also with the developmental stage of D immitis commonly present in the groups at each time point and the number of adult D immitis found at necropsy.

Serial sections of bone and soft tissue from the metacarpophalangeal joints of 2 mature and 2 immature horses were evaluated for substance P immunoreactive sensory nerve fibers. Formalin-fixed specimens were sectioned, either nondemineralized or demineralized with formic acid or EDTA. Rabbit antiserum to substance P (SP) was used in the strep. tavidin-biotin-peraxidase complex method for immunolocalization of SP antigen, and staining with 3,3'- diaminobenzidine was used for permanent identification of SP fibers. Abundant sensory nerve fibers were identified in the joint capsule, synovial membrane subintimal layers, collateral ligaments, suspensory ligament and distal sesamoidean ligament attachments to the sesamoid bones, and the periarticular periosteal layers. Sparse SP-immunoreactive nerve fibers were found in subchondral bone plates of the metacarpus, proximal first phalanx, and dorsal articular surface of the sesamoid bones. Most SP fibers were associated with blood vessels in the small cancellous spaces and haversian canals of the subchondral bone. The deeper marrow spaces contained increased numbers of SP sensory fibers; a few appeared in small groups and as several SP-immunoreactive fibers in a larger nerve. Cortical bone contained only a few SP fibers in the haversian canals. Substance P fibers were not identified in the osteocytic lacunae, canaliculi, or the bony lamellae of the haversian systems of the subchondral bone plate, and its extension to the metaphyseal and diaphyseal cortical bone. Equine metacarpophalangeal joint soft tissues have an abundant sensory nerve supply, similar to that of other species. However, the subchondral bone plate also has sparse sensory nerve fibers, which is a unique finding, and may help explain signs of bone pain associated with disease states of the fetlock.

Serial sections of bone and soft tissue from the metacarpophalangeal joints of 2 mature and 2 immature horses were evaluated for substance P immunoreactive sensory nerve fibers. Formalin-fixed specimens were sectioned, either nondemineralized or demineralized with formic acid or EDTA. Rabbit antiserum to substance P (SP) was used in the strep. tavidin-biotin-peraxidase complex method for immunolocalization of SP antigen, and staining with 3,3'- diaminobenzidine was used for permanent identification of SP fibers. Abundant sensory nerve fibers were identified in the joint capsule, synovial membrane subintimal layers, collateral ligaments, suspensory ligament and distal sesamoidean ligament attachments to the sesamoid bones, and the periarticular periosteal layers. Sparse SP-immunoreactive nerve fibers were found in subchondral bone plates of the metacarpus, proximal first phalanx, and dorsal articular surface of the sesamoid bones. Most SP fibers were associated with blood vessels in the small cancellous spaces and haversian canals of the subchondral bone. The deeper marrow spaces contained increased numbers of SP sensory fibers; a few appeared in small groups and as several SP-immunoreactive fibers in a larger nerve. Cortical bone contained only a few SP fibers in the haversian canals. Substance P fibers were not identified in the osteocytic lacunae, canaliculi, or the bony lamellae of the haversian systems of the subchondral bone plate, and its extension to the metaphyseal and diaphyseal cortical bone. Equine metacarpophalangeal joint soft tissues have an abundant sensory nerve supply, similar to that of other species. However, the subchondral bone plate also has sparse sensory nerve fibers, which is a unique finding, and may help explain signs of bone pain associated with disease states of the fetlock.

Polymorphonuclear neutrophils (PMN) from 4 cows were preincubated (30 minutes, 37 C) in either actinomycin D (100 micrograms/ml) or puromycin (10 micrograms/ml), inhibitors of mRNA transcription and protein translation, or in medium 199. The PMN were incubated for a further 4.5 hours in medium containing 100 U of recombinant bovine interferon-gamma (rboIfn-gamma). The PMN were then incubated with bovine IgG1, IgG2, IgM, or aggregated IgG (aIgG; 4 C, 12 hours) for flow cytometric analysis, using fluoresceinated isotype-specific antibody. The percentage of PMN binding the ligand and the logarithmic mean fluorescent channel (LMFC), an indicator of the amount of receptor (R) expression, were recorded. Competitive inhibition of ligand binding was measured by incubating PMN with fluoresceinated IgG2 in the presence or absence of 100-fold excess of IgG1, IgG2, and aIgG. Activation with rboIfn-gamma induced a 4.5-fold increase in binding of IgG1 and a fivefold increase in LMFC for IgG2. These increases were inhibited by actinomycin D and puromycin. Percentage of PMN binding aIgG decreased after activation by rboIfn-gamma. Interferon-gamma treatment did not affect binding or LMFC of IgM. However, binding of IgM was reduced by treatment with actinomycin D. Binding of fluoresceinated IgG2 was inhibited by unlabeled IgG1, IgG2, and aIgG. Results indicate that bovine PMN Fc receptors (FcR) for IgG1 and IgG2 were rboIfn-gamma inducible, that induction required de novo transcription and translation, that a heterogeneous population of FcR exist on bovine PMN, and that IgG1 and IgG2 share a common FcR. Further, bovine PMN are capable of gene activation and are responsive to changes in their environment, thus being amenable to modulation for effective pathogen destruction.

Polymorphonuclear neutrophils (PMN) from 4 cows were preincubated (30 minutes, 37 C) in either actinomycin D (100 micrograms/ml) or puromycin (10 micrograms/ml), inhibitors of mRNA transcription and protein translation, or in medium 199. The PMN were incubated for a further 4.5 hours in medium containing 100 U of recombinant bovine interferon-gamma (rboIfn-gamma). The PMN were then incubated with bovine IgG1, IgG2, IgM, or aggregated IgG (aIgG; 4 C, 12 hours) for flow cytometric analysis, using fluoresceinated isotype-specific antibody. The percentage of PMN binding the ligand and the logarithmic mean fluorescent channel (LMFC), an indicator of the amount of receptor (R) expression, were recorded. Competitive inhibition of ligand binding was measured by incubating PMN with fluoresceinated IgG2 in the presence or absence of 100-fold excess of IgG1, IgG2, and aIgG. Activation with rboIfn-gamma induced a 4.5-fold increase in binding of IgG1 and a fivefold increase in LMFC for IgG2. These increases were inhibited by actinomycin D and puromycin. Percentage of PMN binding aIgG decreased after activation by rboIfn-gamma. Interferon-gamma treatment did not affect binding or LMFC of IgM. However, binding of IgM was reduced by treatment with actinomycin D. Binding of fluoresceinated IgG2 was inhibited by unlabeled IgG1, IgG2, and aIgG. Results indicate that bovine PMN Fc receptors (FcR) for IgG1 and IgG2 were rboIfn-gamma inducible, that induction required de novo transcription and translation, that a heterogeneous population of FcR exist on bovine PMN, and that IgG1 and IgG2 share a common FcR. Further, bovine PMN are capable of gene activation and are responsive to changes in their environment, thus being amenable to modulation for effective pathogen destruction.