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Abstract
Summary
This study was conducted on White Leghorn chicken. Specific pathogen free one day chicks were kindly provided from Goto Chick Company Gifu, Japan. The study was approved by Gifu University, Animal Care and Use and it was in accordance with Japanese Department of Agriculture guidelines
1. Preference behavior to capsaicin in chicken
A) Preference to capsaicin in chicken
• In drinking bottles preference test starting from 1day post-hatching, chicks showed equal preference to capsaicin (CAP; 10 ppm), vehicle or plain drinking water during the first week.
• In the 2nd week; chicken preference shifted towards capsaicin and at 3rd week chicken preferred completely capsaicin and refused either vehicle or plain drinking water.
B) Induction of Preference to capsaicin in chicken
• Pretreatment with CAP (10mg/kg, BW,IV) daily from the 1st day to the 3rd day age induced preference to capsaicin than vehicle or drinking water after 3 weeks from treatment.
2. Thermoregulatory response to capsaicin
1) At 3 days chicks injection of capsaicin (10mg / kg,BW,IV) caused fall in colonic temperature (Tc) (hypothermia down to 40.1±0.18 at 1.5 hours post-injection) that continued over 4 hours followed by thermoregulatory response started from 5 hours onwards (regaining Tc to normal 40.8 ± 0.02).
2) At 4 days age when the same chicks were injected with CAP (10mg/kg, BW,IV.) these chicks developed delayed onset of hypothermia (down to 40.38 ± 0.2°C, P < 0.05) that was of short duration (about 1 hour) and accompanied by thermoregulatory response 6 hours after injection.
3) While at 5 days chicks, third CAP (10 mg/kg,IV) application did not cause any significant changes in Tc. The differences in the effect of CAP on Tc among 3, 4, and 5 days were not related to the age factor. As when CAP (10mg/ kg,BW,IV) was injected in 5 days chicks for the first time induced significant hypothermia that continued for 1 hour,
4) On the other hand, CAP (10 mg / kg,BW,IV) hypothermic effect did not appear in chicks over 10 days. In the later case the differences might be attributed to age factor.
3. Effect of capsaicin on behavioral and thermoregulatory response to heat in chicken
1) The percent of escape within 50 min from heat chamber was used to indicate the effect of CAP on accommodation to high ambient temperature (40°C). Chicks 7- days pretreated with CAP (10 mg / kg,BW,IV) showed the best adaptation indicated by lowest escape to hot environment ((40 ± 12) %) followed by the non treated control (78 ± 6 %) while CAP (100 mg/ kg,BW,IV) pre-injected group was the worst (96 ± 4 %).
2) Also 7- days pretreated chicks with CAP (10 mg / kg,BW,IV) showed better thermoregulation to heat (37 °C ambient temperature) indicated by lower Tc 41.7 ± 0.1°C and thermal index 1.6 °C after heat exposure. While the Tc was 42.1± 0.1°C and thermal index was 5.3 °C in non treated group. At the same time it was 1.1 °C for control group kept at ambient temperature 25°C.
4. Effect of capsaicin on nociceptive response to ammonia solution
CAP (10 mg /kg BW,IV) pretreatment decreased the reaction time (sec) and number of eye wips (30sec) to topical instillation of 0.1%NH4OH but not to 1% NH4 OH solution in 4 and 10 days age chicks.
A) In 4 day chicks CAP (10 mg / kg,BW,IV) pretreatment decreased reaction time (11 ± 2.0 sec) when compared to non treated control (76.9 ± 11.2 sec) and number of eye wips (2.5 ± 2.0/30sec) when compared to (13 ± 2.33/30sec) in non treated control chicks.
B) Also in 10 day chicks CAP (10 mg/kg,BW,IV, pretreatment 7 days before) reduced the reaction time to (2.5 ± 0.50 sec.) compared to untreated control ‘22 ± 3.5 sec.) and number of eye wips was also 2 ± 0.53/30sec when compared to (17 ± 5.4 /30sec) in control chicks.
5. Lipopolysaccharide - induced thermoregulatory response in chicken
1) LPS induced changes in Tc were dose dependent. They ranged from fever at 2 and 5 mg/kg BW.IV to hypothermia at 10 mg/kg,BW,IV. LPS induced changes in Tc were also age dependent.
2) At 4 days chicks LPS (2 mg/kg BW.IV) induced monophasic fever (about 1°C) , while in 10 days chicks it induced polyphasic fever with rises of Tc reaching peaks; (42.42 °C) at 2.5, (42.15°C) at 4.5 and (42.41 °C) at 6 hours respectively after LPS injection .
3) whereas LPS (10 mg/kg BW.IV) induced hypothermia varied from hypothermic shock (37.75 ± 0.5°C) in 4 days chicks to prolonged hypothermia (40.04 ± 0.29) at 4.5 hours that continued for about 7 hours in 10 days chicks.
4) In five weeks chicken, LPS (2 mg/ kg,BW,IV) induced polyphasic pattern; initial hypothermia about 1.1°C directly after injection followed by two rises in Tc (42.10± 0.2°C) at 2.5 hours and (42.4± 0.1°C) at 7 hours after LPS injection.
5) While LPS (10 mg/ kg,BW,IV) at 5 weeks did not induced prolonged hypothermia but induced only initial hypothermia (39.95 °C) followed by two transient febrile phases of intensities (about 42.3 °C) at 3.5 and 42.4°C at 6.5 hours after LPS injection respectively.
6. Capsaicin pretreated prevented LPS -induced hypothermia and septic shock
At 4 day age CAP (10mg/kg,BW,IV) pretreated chicks showed rapid thermoregulation (Tc; 40.2 ± 0.2 °C) started 2 hr after LPS (10mg/kg,BW,IV) injection and rescued from LPS induced hypothermic shock (survivability; 100%) within 5 hours time. Also in 10 day chicks CAP pretreatment prevented LPS- induced hypothermia and chicks started to thermoregulate (40.7± 0.1°C) within 5.5 hour time.
7. Capsaicin pretreatment prevented cold -induced hypothermia and hypothermic collapse
In 4 day chicks acute cold stress (8 °C) induced hypothermic collapse (Tc down to 38.65 ± 0.06°C) at 4.5 hours and 90.9% mortality within 5 hours cold exposure. However, CAP pretreatment showed better thermoregulation (40.55 ± 0.2 °C) and rescued from cold induced hypothermic collapse (survivability; 83.3%) within the 5 hours of cold exposure.
Also in 10 days chicks, acute cold exposure for 10 hours induced prolonged hypothermia (40.18 ± 0.1°C) at 1 hour and continued for 8 hours after cold exposure. While CAP in 10 days chicks (7 days CAP pretreated) showed rapid thermoregulation started 3.5 hour after cold exposure and continued for 8 hours of cold exposure.
8. Capsaicin pretreated prevented LPS induced fever
1) At 5 days age LPS (2 mg /kg,BW,IP) induced slight and transient decrement in Tc followed by an increment in temperature(fever) that started 2 hours post injection, reached the peak (42.1± 0.1°C) at 2.5 hours post injection and continued for about 2.5 hour. This LPS induced febrile response was highly significantly attenuated (41.2 ± 0.1°C) in CAP (10 mg/ kg,BW,IV) pretreated chicks.
2) In 10 days chicks LPS (2 mg /kg,BW,IP) induced polyphsic fever. It composed of initial decrement (40± 0.14°C) in Tc followed by three peaks of fever; 42.23± 0.21° °C, 42.2 ± 0.24 °C and 42.4 ± 0.15° °C at 3 hours, 6.5 hours and 8 hours after LPS injection respectively. The onset of the response (the entire first phase and a part of the second phase) was significantly attenuated at 3 hours after LPS, (Tc = 41.0 ± 0.11°C) in CAP-pretreated chicks.
9. Capsaicin pretreatment prevented LPS induced Sickness behaviors in chicken
A) LPS induced anorexia
Injection of LPS (2mg/kg,BW,IP) induced abrupt stop of eating (anorexia) starting directly after injection.
1- Capsaicin pretreatment did not block LPS induced anorexia in 5 days chicks within 8 hours after LPS injection.
1- Capsaicin pretreatment increased significantly the food intake in 10 day chicken at 8 hours (1.2 ± 0.13g /5min /chick), (Fig.23 A, B&C) and approached the control (1.43 ± 0.06 g/chick/5min) .
2- Also in 5 week chicks at 8 hours after LPS injection, capsaicin increased the intake rate (3.1 ± 0.2 g / chick/ 5min) and nearly exceeded the control (2.97 ± 0.12 g/ 5min / chick).
B) LPS induced somnolence
CAP pretreatment attenuated LPS induced somnolence unexceptionally of the age effect during 8 hours after LPS injection.
1- In non treated 5 day chicks LPS induced somnolence (210 and 245 (sec/5min/chick) at 4 and 8 hours post injection respectively). While in CAP pretreated chicks it was reduced to 79 and 113 (sec/5min/chick) at 4 and 8 hours post injection respectively.
2- In 10 days chicks, somnolence time in LPS/CAP treated chicks at 4 hours was 101 (sec/5min/chick) compared to180 (sec/5min/chick) in LPS injected chicks. Similarly, CAP was more effective on LPS induced somnolence after 8 hour of injection (LPS/CAP = 67 sec/5min, compared to LPS 130 sec/5min/ chick)
Also CAP pretreatment was effective after 2 hours from LPS injection in 5 weeks chicks (LPS/CAP = 187 sec/5min, compared to LPS 270 sec/5min/ chick).
C) LPS induced adipsia
Directly after injection of LPS drinking was stopped in all chicks regardless of age. However CAP (10 mg/kg,BW,IV) pretreatment reversed LPS induced adipsia in all age groups during the first 4 hours after injection.
10. Modulatory role of capsaicin to LPS- induced innate and adaptive immunity in chicken
10.1. Effect of CAP on LPS induced NO production and iNOS gene expression
1.Three hours after LPS (2mg/ kg,BW,IP) injection nitrite production was increased to 345 , 200 and 250 (nmol/mg) in liver, Lung and hypothalamus respectively. In 10 day chicks LPS increased nitrite up to 348, 395 and 220 (nmol/mg) in liver, lung and brain respectively. While, in both 4 and 10 days chicks, CAP (10mg/kg,BW,IV) pretreatment significantly (P < 0.05) attenuated LPS induced nitrite production in all groups
2. In all tested organs (liver, lung and brain) of both age groups iNOS expression was elevated after LPS injection. While in all organs except hypothalamus, CAP pretreatment significantly inhibited LPS induced iNOS gene expression in both age groups of chicks.
10.2. Effect of CAP on LPS induced MHC class II genes expression
1. LPS (2mg/ kg,BW,IP) increased MHC class II complex (BL-complex) genes expression in bursa of Fabricius. Twelve hours after LPS injection, the inducible levels of MHC II estimated by relative fold increase of (BL-α) / beta actin were 0.98 and 0.74 in 4 and 10 days chicks respectively. CAP pretreatment significantly (P< 0.05) inhibited LPS induced BL- α gene expression in both 4 and 10 days chicks.