Stress and Leukocytes Home

If you've been reading the tables in the previous pages and noticed that for some well-studied species, the percentage of lymphocytes and neutrophils differ greatly between studies, it's likely because of the stress levels of the animals being studied. The effects of stress on circulating white blood cell distributions has been experimentally studied in a number of animal species over several decades (reviewed in Davis et al. 2008). Below is a table that summarizes this literature (at least the papers the author knows of). In most of these studies, animals were injected with stress hormones - corticosterone, cortisol, dexamethasone or ACTH (which induces cort production) to mimic a stress response, though some involved manipulation of endogenous stress hormones, by way of physical restraint, transport, amputation (newts), temperature extremes, or simply capturing wild animals, which they perceive as stressful.

Last Updated Dec 30 2011

Source

Journal

Subjects

Method

Result

Salak-Johnson et al. 1996

Journal of Animal Science

Pigs

Injections of ACTH and Cortisol

ACTH caused increases in neutrophils, decreases in lymphocytes, rise in N/L, cortisol caused decreases in lymphocytes

Bilandžič et al. 2006

Veterinary Research Communications

Boars

Injections with ACTH over 3 days

Significant increases in percentage of neutrophils, decreases in percentage of lymphocytes, significant rise in N/L ratio

Burguez et al. 1983

Equine Veterinary Journal

Horses

Injections with cortisol, sampled multiple times after

Significant increases in N/L ratio after 2 hours in foals, after 4 hours in adults

Rossdale et al. 1982

Equine Veterinary Journal

Horses

Injections with ACTH

Significant increases in N/L ratio after 4 hours in adults

Frank et al. 2006

Canadian Veterinary Journal

Dogs

Sampled before and after transport

Nonsignificant increase in N/L ratio after transport

Kock et al. 1999

Journal of Zoo and Wildlife Medicine

Rhinoceros

Sampled before and after transport

Significant increase in neutrophils, decrease in lymphocytes and eosinophils

Reidarson and McBain 1999

Journal of Zoo and Wildlife Medicine

Dolphins

Single oral dose of dexamethasome (glucocorticoid), sampled multiple times after

Significant neutrophilia, lymphopenia and eosinopenia after 24 hours, returned to normal 48 hours later

McLaren et al. 2003 Experimental Physiology Badgers Compared animals transported to those not transported Significant increase in percentage of neutrophils, significant decrease in lymphocytes, increase in N/L ratio in transported animals

Kranendonk et al. 2005

American Journal of Veterinary Research

Female Pigs

Twice-daily oral administration of hydrocortisone acetate from 7-11 weeks of gestation

N/L ratios significantly higher in treated pigs than in controls

Anderson et al. 1999

Veterinary Research Communications

Cows

Injections of dexamethasome

Injected animals had neutrophilia, lymphopenia, eosinopenia, increased N/L ratio, monocytosis

Trottier et al. 2008

PNAS

Mice

Implantation with cort tablets, blood sampled 3 days later

Elevated neutrophil counts, lowered lymphocyte counts in blood

Montane et al. 2003

Journal of Wildlife Diseases

Deer

Deer were captured, samples obtained at capture and at 1, 2 and 3 hours after

Decrease in lymphocyte count

Dhabhar et al. 1994

Brain, Behavior and Immunity

Mice

Physical restraint for 1 hour

Restrained mice had reduced relative numbers of lymphocytes, increased percentage of neutrophils compared to controls (non-restrained mice)

López-Olvera et al. 2007

Canadian Journal of Veterinary Research

Chamois

Capture and physical restraint, sampling each hour thereafter for 3 hours

Absolute neutrophil counts increased, lymphocyte counts decreased

McGlone et al. 1993

Journal of Animal Science

Pigs

Sampled before and after 4hr transport

Numbers of neutrophils increased, lymphocytes decreased, N/L ratio increased after shipping

Morrow-Tesch et al. 1993

Psychoneuroendocrinology

Macaques

Physical restraint for 3 hours, sampling hourly

Percentage of neutrophils increased, percentage of lymphocytes decreased steadily over 3 hours

Aguirre et al. 1995

Physiological Zoology

Sea Turtles

Capture of wild individuals, sampling at 2 min, 1 hr, 4 hr, and 24 hr, H/L and cort levels monitored

Cort increased after 1 hr, remained high, H/L did not increase until 24 hrs later

Johansson-Sjobeck et al. 1978 Comparative Biochemistry and Physiology Eels Daily injections with cortisol over 14 days, measuring WBCs at day 1, 4, 14 Decrease in number of lymphocytes after 4 days , increase in number of neutrophils after 14 days
Balabanova et al. 2009 Inland Water Biology Carp Injection with dexametazone, measuring WBCs at 1,3,7,14 and 21d after Reduction in percentage of lymphocytes, increase in percentage of neutrophils

Bennett and Reap 1978

Journal of Comparative Physiology

Eastern Newts

Exposure to constant light

Relative increase in neutrophil numbers (relative to control animals under normal light cycles), decrease in relative lymphocytes

Bennett and Alspaugh 1964

Virginia Journal of Science

Leopard frogs

Single injection of hydrocortisone acetate sampled before and 72 and 144 hrs after

Increase in proportion of neutrophils at all time periods (relative to control animals), decrease in lymphocytes at all time periods

Bennett and Newell 1965

Virginia Journal of Science

Leopard frogs

Single injection of hydrocortisone acetate sampled before and 12, 24, 48 hrs after

Increase in proportion of neutrophils at all time periods (relative to control animals), decrease in lymphocytes at all time periods

Bennett and Harbottle 1968

Biological Bulletin

Larval bullfrogs

Single injection of hydrocortisone acetate sampled before and 48 hrs after

Increase in proportion of neutrophils (relative to control animals), decrease in lymphocytes

Bennett and Johnson 1973

Journal of Comparative Physiology

Eastern Newts

Single injection of ACTH, or immersion in NaCl solution for 2 hrs, sampled before and 24 and 48 hrs after

Increase in proportion of neutrophils (relative to control animals) in both treatments at all times, decrease in lymphocytes also

Bennett and Daigle 1983

Journal of Comparative Physiology

Eastern Newts

Exposure to 8d of low (3C) or high (21C) temps

Increase in proportion of neutrophils in low and high temp treatments relative to animals in control (11C temp), decrease in lymphocytes also

Bennett et al. 1972

Journal of Comparative Physiology

Eastern Newts

Single injection of ACTH, sampled before and 48, 72hrs after

Increase in proportion of neutrophils, decreases in proportion of lymphocytes (relative to controls) at 72 hrs (not 48)

Bennett 1986

Journal of Comparative Physiology

Eastern Newts

Amputation of forelimb, sampling 8, 15, 22 days later

Relative increase in neutrophils, decrease in lymphocytes at all time periods

Reed and Bennett 1973

Journal of Comparative Physiology

Eastern Newts

Single injection of hydrocortisone acetate at 4 different times of day, sampled after 72 hrs

Increase in relative neutrophil count, decrease in relative lymphocyte count at all times, most pronounced late in day, least early in day

Bennett et al. 1974

Journal of Comparative Physiology

Eastern Newts

Single injection of ACTH at 8 different times of day, sampled after 72 hrs

Increase in relative neutrophil count, decrease in relative lymphocyte count at all times

Davis and Maerz 2010

International Journal of Zoology

mole salamanders

Injections with corticosterone to wild caught salamanders, sampling 24 hrs later

Increase in N/L ratio over sham and control groups

Davis and Maerz 2011 Journal of Herpetology marbled salamanders, leopard frogs Capture from ponds, chased by hand for 1 min in individual containers, blood sampled 24 hr later for leukocyte counts Threefold increase in average N/L ratios in both species compared to baseline samples at time of capture
Narayan and Hero 2011 Australian Journal of Zoology Fifian ground frogs Capture and transport (6 hr) followed by captivity (15d), leukocyte data and cort levels monitored N/L ratios increased after 6hr transport and after 15 days of captivity, returned to baseline after 25 days. N/L was positively corellated with cort at all time periods

Lance and Elsey 1999

Journal of Experimental Zoology

Alligators

Physical restraint for 48 hours, samples taken throughout

Increase in percentage of heterophils, decrease in percentage of lymphocytes, rise in H/L ratio over entire period

Altan et al. 2000

Turkish Journal of Veterinary Animal Science

Chickens

Exposure to increased temperature for 2 hr

Increased proportions of heterophils and basophils, decreased proportions of lymphocytes and monocytes, increased overall H/L ratio

Bedáňová et al. 2007

Acta Vet BRNO

Chickens

Suspension from shackles (mimicking processing plant conditions) for 2 or 5 min, sampled 20h later

Increase in heterophil counts compared to control birds in both groups, increase in basophils in 5min group.

Zulkifli et al. 1999

Poultry Science

Chicks

Capture and handling, sampled 20h later

Increase in H/L ratio in handled chicks compared to controls

McFarlane et al. 1989

Poultry Science

Chicks

Intermittent electric shock, heat stress and aerial ammonia for 7 days

H/L ratio increased with each single treatment, increased further with multiple stressors

Aengwanich and Chinrasri 2003

Songklanakarin J. Sci. Technol.

Quail

Oral doses of dexamethasone for 4 days, sampling at day 1,3,7,10,14

Percentage of heterophils increased, lymphocytes decreased on day 3, returned to normal on day 10

Gross and Siegel 1983 **

Avian Diseases

Chickens

Various doses of corticosterone mixed with food for 1 day, sampled day 2

H/L ratios increased in all cort treatments, also showed a linear relationship between cort dose and H/L ratio

Maxwell 1993

World’s Poultry Science Journal

Mostly Chickens

Multiple stressors

A review of the early literature (from the 1980s) on the avian leukocyte response to stress

*Neutrophilia = increase in circulating neutrophils, Lymphopenia = decrease in circulating lymphocytes

** The Gross and Siegel (1983) paper is widely-recognized as the first publication on the subject of stress and leukocytes in birds, and deserves further comment here. Gross was from the College of Veterinary Medicine at Virginia Tech, while Siegel was at the USDA Southeast Poultry Research Lab in Athens, GA. Their paper described for the first time the tight relationship between stress hormones and H/L ratios in birds (chickens). They conducted a suite of experiments, stressing birds in various ways, then measuring H/L ratios. The most important experiment was also the simplest and most elegant; they mixed corticosterone into the chicken's feed for one day at varying doses and compared H/L ratios, and plasma levels of corticosterone at the end of the day. They found that birds fed corticosterone had a significantly higher H/L ratio than control birds, but more importantly there was a positive, linear relationship between the cort dose administered and mean H/L ratios per cage (each cage had 10 birds). Interestingly, the relationship between initial dose and plasma levels of corticosterone was much weaker (right graph).

Below are graphs of the data presented by Gross and Siegel (Table 2 in the paper). Each point is the mean of 10 birds.

 

The authors concluded that H/L ratios represent a reliable indicator of long-term stress, while plasma levels of corticosterone could be considered better-suited for indexing short-term stress.

 

Complete Reference List for Citations in Table

Aengwanich, W., and O. Chinrasri. 2003. Effect of dexamethasone on differential white blood cell counts and heterophil/lymphocyte ratio in Japanese quails (Coturnix coturnix japonica). Songklanakarin J. Sci. Technol. 25: 183-189.

Aguirre, A. A., G. H. Balazs, T. R. Spraker, and T. S. Gross. 1995. Adrenal and hematological responses to stress in juvenile green turtles (Chelonia mydas) with and without fibropapillomas. Physiological Zoology 68: 831-854.

Altan, O., A. Altan, M. Cabuk, and H. Bayraktar. 2000. Effects of heat stress on some blood parameters in broilers. Turkish Journal of Veterinary Animal Science 24: 145-148.

Anderson, B.H., D.L. Watson, and I.G. Colditz. 1999. The effect of dexamethasone on some immunological parameters in cattle. Veterinary Research Communications 23: 399-413.

Balabanova, L. V., D. V. Mikryakov, and V. R. Mikryakov. 2009. Response of common carp (Cyprinus carpio L.) leucocytes to hormone-induced stress. Inland Water Biology 2: 86-88.

Bedáňová, I., E. Voslářová, V. Večerek, V. Pištěková, and P. Chloupek. 2007. Haematological profile of broiler chickens under acute stress due to shackling. Acta Veterinaria Brno 76: 129-135.

Bennett, M.F. 1986. Stress and changes in the blood of newts, Notophthalmus viridescens, during early regeneration. Journal of Comparative Physiology A 159: 823-826.

Bennett, M.F., and J.K. Alspaugh. 1964. Some changes in the blood of frogs following administration of hydrocortisone. The Virginia Journal of Science 15: 76-79.

Bennett, M.F., and N.C. Newell. 1965. A further study of the effects of hydrocortisone on the blood of frogs. The Virginia Journal of Science 16: 128-130.

Bennett, M.F., and J.A. Harbottle. 1968. The effects of hydrocortisone on the blood of tadpoles and frogs, Rana catesbeiana. Biological Bulletin 135: 92-95.

Bennett, M.F., and A.O. Johnson. 1973. Osmotic stress, ACTH and the white blood cell picture in newts, Notophthalmus viridescens. Journal of Comparative Physiology A 82: 333-338.

Bennett, M.F., and L.E. Reap. 1978. Photoperiod, stress and the distribution of leukocytes in the peripheral blood of Notophthalmus viridescens. Journal of Comparative Physiology A 125: 205-207.

Bennett, M.F., and K.R. Daigle. 1983. Temperature, stress and the distribution of leukocytes in red-spotted newts, Notophthalmus viridescens. Journal of Comparative Physiology A 153: 81-83.

Bennett, M.F., C. Reed, and R.L. Roden. 1974. Circadian changes in responses of newts, Notophthalmus viridescens, to ACTH. Journal of Comparative Physiology A 89: 287-291.

Bennett, M.F., C.A. Gaudio, A.O. Johnson, and J.H. Spisso. 1972. Changes in the blood of newts, Notophthalmus viridescens, following administration of hydrocortisone. Journal of Comparative Physiology A 80: 233-237.

Bilandžič, N., M. Žurič, M. Lojkič, B. Šimič, D. Milić, and I. Barač. 2006. Cortisol and immune measures in boars exposed to three-day administration of exogenous adrenocorticotropic hormone. Veterinary Research Communications 30: 433-444.

Burguez, P.N., J. Ousey, R.S.G. Cash, and P.D. Rossdale. 1983. Changes in blood neutrophil and lymphocyte counts following administration of cortisol to horses and foals. Equine Veterinary Journal 15: 58-60.

Davis, A.K. and J.C. Maerz. 2010. Effects of exogenous corticosterone on circulating leukocytes of a salamander (Ambystoma talpoideum) with unusually abundant eosinophils. International Journal of Zoology. DOI:10.1155/2010/735937

Davis, A. K. & Maerz, J. C. 2011 Assessing stress levels of captive-reared amphibians with hematological data: implications for conservation initiatives. Journal of Herpetology 45, 40-44.

Dhabhar, F.S., A.H. Miller, M. Stein, B.S. McEwen, and R.L. Spencer. 1994. Diurnal and acute stress-induced changes in distribution of peripheral blood leukocyte subpopulations. Brain Behavior and Immunity 8: 66-79.

Frank, D., A. Gauthier, and R. Bergeron. 2006. Placebo-controlled double-blind clomipramine trial for the treatment of anxiety or fear in beagles during ground transport. Canadian Veterinary Journal-Revue Veterinaire Canadienne 47: 1102-1108.

Gross, W.B., and H.S. Siegel. 1983. Evaluation of the heterophil/lymphocyte ratio as a measure of stress in chickens. Avian Diseases 27: 972-979.

Johansson-Sjobeck, M. L., et al. (1978). Hematological effects of cortisol in european eel, anguilla anguilla l. Comparative Biochemistry and Physiology a-Physiology, 60: 165-168.

Kock, R.A., S.R.O. Mihok, J. Wambua, J. Mwanzia, and K. Saigawa. 1999. Effects of translocation on hematologic parameters of free-ranging black rhinoceros (Diceros bicornis michaeli) in Kenya. Journal of Zoo and Wildlife Medicine 30: 389-396.

Kranendonk, G., H. Hopster, F. van Eerdenburg, K. van Reenen, M. Fillerup, J. de Groot, M. Korte, and M. Taverne. 2005. Evaluation of oral administration of cortisol as a model for prenatal stress in pregnant sows. American Journal of Veterinary Research 66: 780-790.

Lance, V.A., and R.M. Elsey. 1999. Plasma catecholamines and plasma corticosterone following restraint stress in juvenile alligators. Journal of Experimental Zoology 283: 559-565.

López-Olvera, J.R., I. Marco, J. Montané, E. Casas-Díaz, and S. Lavín. 2007. Effects of acepromazine on the stress response in Southern chamois (Rupicapra pyrenaica) captured by means of drive-nets. Canadian Journal of Veterinary Research 71: 41-51.

Maxwell, M.H. 1993. Avian blood leukocyte responses to stress. Worlds Poultry Science Journal 49: 34-43.

McFarlane, J.M., S.E. Curtis, J. Simon, and O.A. Izquierdo. 1989. Multiple concurrent stressors in chicks 2. Effects on hematologic, body composition, and pathologic traits. Poultry Science 68: 510-521.

McGlone, J.J., J.L. Salak, E.A. Lumpkin, R.I. Nicholson, M. Gibson, and R.L. Norman. 1993. Shipping stress and social-status effects on pig performance, plasma cortisol, natural killer cell activity, and leukocyte numbers. Journal of Animal Science 71: 888-896.

McLaren, G. W., D. W. Macdonald, C. Georgiou, F. Mathews, C. Newman, and R. Mian. 2003. Leukocyte coping capacity: a novel technique for measuring the stress response in vertebrates. Experimental Physiology 88: 541-546.

Montane, J., I. Marco, J. Lopez-Olvera, D. Perpinan, X. Manteca, and S. Lavin. 2003. Effects of acepromazine on capture stress in roe deer (Capreolus capreolus). Journal of Wildlife Diseases 39: 375-386.

Morici, L.A., R.M. Elsey, and V.A. Lance. 1997. Effects of long-term corticosterone implants on growth and immune function in juvenile alligators, Alligator mississippiensis. Journal of Experimental Zoology 279: 156-162.

Morrow-Tesch, J.L., J.J. McGlone, and R.L. Norman. 1993. Consequences of restraint stress on natural killer cell activity, behavior, and hormone levels in Rhesus Macaques (Macaca mulatta). Psychoneuroendocrinology 18: 383-395.

Narayan, E., and J. M. Hero. 2011. Urinary corticosterone responses and haematological stress indicators in the endangered Fijian ground frog (Platymantis vitiana) during transportation and captivity. Australian Journal of Zoology 59: 79-85.

Reed, C., and M.F. Bennett. 1973. Circadian effects of hydrocortisone on the blood of the newt, Notophthalmus viridescens. Journal of Comparative Physiology A 86: 59-63.

Reidarson, T.H., and J.F. McBain. 1999. Hematologic, biochemical, and endocrine effects of dexamethasone on bottlenose dolphins (Tursiops truncatus). Journal of Zoo and Wildlife Medicine 30: 310-312.

Rossdale, P.D., P.N. Burguez, and R.S.G. Cash. 1982. Changes in blood neutrophil/lymphocyte ratio related to adrenocortical function in the horse. Equine Veterinary Journal 14: 293-298.

Salak-Johnson, J.L., J.J. McGlone, and R.L. Norman. 1996. In vivo glucocorticoid effects on procine natural killer cell activity and circulating leukocytes. Journal of Animal Science 74: 584-592.

Trottier, M.D., M.M. Newsted, L.E. King, and P.J. Fraker. 2008. Natural glucocorticoids induce expansion of all developmental stages of murine bone marrow granulocytes without inhibiting function. Proceedings of the National Academy of Sciences of the United States of America 105: 2028-2033.

Zulkifli, I., M.T.C. Norma, C.H. Chong, and T.C. Loh. 1999. Heterophil to lymphocyte ratio and tonic immobility reactions to preslaughter handling in broiler chickens treated with acorbic acid. Poultry Science 79: 402-406.