26. Prediction of imminent death in mice used for longevityresearchM.A. Ray, N.A. Johnston, S.J. Verhulst, L.A. Toth

Southern Illinois University School of Medicine, Springfield, IL 62794,USA

Background: Mice and other species used in longevity represent asignificant investment in research time and other resources. Maxi-mal utilization of these animals as sources of data would beenhanced if objective accurate markers for imminent death couldbe established to permit timely euthanasia. However, despite theanticipated increase in health problems as aging progresses, manymice are found dead in the cage without obvious antemortem signsof illness. Our goal in this study was to identify objective criteria thatcould be used to predict imminent death in aged mice. If successful,application of such markers would allow planned euthanasia ofthese valuable animals, permitting their use for measurements thatwould otherwise be impossible. Methods: A total of 105 eight-month-old ICR mice (retired breeders; 58 females and 47 males;Harlan) were subcutaneously implanted with an identification chipthat also allowed remote measurement of body temperature. Malemice were housed individually to avoid injury due to fighting,whereas female mice were assigned to either group (10 mice percage) or individual housing. Mice were evaluated weekly for bodytemperature, weight, and visible signs of illness or aging, includinggeneral body condition, respiratory rate and character, dehydration,posture, hair coat condition, spontaneous ambulation, and responseto manipulation. Eighty-one of the mice were monitored until spon-taneous death occurred, whereas 24 underwent euthanasia based onclinical signs. Clinical signs that were used to signal implementationof euthanasia were inability or unwillingness to walk, lack of overtresponse to manipulation, large or ulcerated tumors, and/or palpablehypothermia. Results: Housing and gender did not significantly influ-ence outcome measures. Therefore, data from all mice was allocatedinto two groups – spontaneous death versus euthanasia – for subse-quent analysis. Mice that underwent euthanasia (n = 24) were killedat a mean age of 25.3 ± 1.3 months, whereas the mean age at deathamong mice that died spontaneously (n = 74) was 24.5 ± 0.7 months.Data from both groups (i.e., spontaneous death and euthanasia) wereanalyzed to identify clinical signs that preceded death. Weight losswas the most common and earliest sign. Mice lost approximately20% of their body weight during the 3 month period that precededdeath, with about half of that loss occurring precipitously duringthe last week of life. Body temperatures were generally stable untilthe last week of life, but then fell by over 1 �C. Slow and/or laboredrespiration was observed in 43% of mice within 24 h of death. Thus,monitoring body weight as a preliminary marker and using hypo-thermia. respiratory difficulty, and/or precipitous weight loss as pen-ultimate markers appears to allow prediction of death to within oneweek. Among the mice used in this study, one week represents onaverage about 1% of the median life span. Closer evaluation ofhigh-risk mice could potentially allow even more accurate predic-tion. Conclusion: Mice that demonstrate two or more of the following

physical indicators can accurately be predicted to die within a max-imum of one week: precipitous decrease in body weight (>10% loss),decreased body temperature (>1 �C decrease), or slow or labored res-piration. Euthanasia of mice based on these criteria may allow col-lection of tissues for subsequent analysis prior to spontaneousdeath of the animal without significantly impacting longevityoutcomes.

doi:10.1016/j.exger.2008.08.040

27. DCNP1 up-regulates corticotropin-releasing hormone geneexpression and may play a role in depressionT. Zhou a,b, S. Wang b, W. Kamphuis a, G. Wang b, J. Zhou b, D.K.Swaab a

a Netherlands Institute for Neuroscience, Amsterdam, The Netherlandsb Hefei National Laboratory for Physical Sciences at Microscale andDepartment of Neurobiology and Biophysics, Life Science School,University of Science and Technology of China, Hefei, Anhui, PR China

Dendritic cell nuclear protein1 (DCNP1) is a newly found proteinthat may be a novel candidate gene for major depression. Recentlypolymorphisms in DCNP1 are reported to have a relationship withdepression. However, the distribution of DCNP1 in the human brainand the way it gets involved in the etiology of depression stillremained unknown. Corticotropin-releasing hormone (CRH) plays apivotal role in depression as a key mediator of the hypothalamic-pituitary-adrenal (HPA) axis that is the final common pathway ofthe signs and symptoms of depression. Therefore we study the rela-tionship between DCNP1 and the HPA-axis and in particular with itsCRH neurons. In the present study, with real-time PCR technique wefound for the first time that DCNP1 is widely distributed in differenthuman brain areas. By this technique we also, showed that themRNA level of DCNP1 increased significantly (p = 0.004) in thelaser-dissected paraventricular nucleus of depressed patients ascompared to controls. In addition, DCNP1 was found to significantly(p = 0.005) up-regulate CRH gene expression in the co-transfectionexperiment and luciferase assay. Further more, chromatin immuno-precitation showed DCNP1 binding to the CRH transcript promoterin the nucleus. Our primary results clarify the basal molecular mech-anism of DCNP1 action on the HPA-axis and suggest a role of thisprotein in depression. Acknowledgments: We are indebted to theNetherlands Brain Bank at the Netherlands Institute for Neurosci-ence for providing us with the brain material and patient informa-tion. This investigation was supported by the China ExchangeProgramme of the Royal Netherlands Academy of Arts and Sciences(KNAW) and the Natural Science Foundation of China (project08CDP012).

doi:10.1016/j.exger.2008.08.041

134 Abstracts / Experimental Gerontology 44 (2009) 126–134