Brain proteome analysis of mice selectively bred for either high or low anxiety-related behavior revealed quantitative and qualitative protein expression differences. The enzyme glyoxalase-I was consistently expressed to a higher extent in low anxiety as compared with high anxiety mice in several brain areas. The same phenotype-dependent difference was also found in red blood cells with normal and cross-mated animals showing intermediate expression profiles of glyoxalase-I. Another protein that showed a different mobility during two-dimensional gel electrophoresis was identified as enolase phosphatase. The presence of both protein markers in red or white blood cells, respectively, creates the opportunity to screen for their expression in clinical blood specimens from patients suffering from anxiety. Biomarkers are measures of biological parameters of disease that also can predict which new molecular entities will be effective and safe in treating patients (1). We have focused our biomarker discovery efforts on the analysis of mouse models (2) to avoid the interindividual differences of human specimens that often result in a low biomarker signal to background noise ratio.The manifestation of anxiety in a number of psychiatric disorders such as generalized anxiety disorder, depression, panic attacks, phobias, obsessive-compulsive disorders, and post-traumatic stress disorder (3) highlights the importance of gaining a better understanding of associated reliable biomarkers in proper animal models. An animal model to study behavioral, neuroendocrine, and genetic concomitants of trait anxiety including psychopathology should represent a good approximation to score symptoms of anxiety disorders and possibly comorbid depression (4 -6). To avoid interstrain comparisons, which are likely to reveal differences in more than just anxiety-related indices, we have been using intrastrain breeding approaches to focus on particular traits, including anxiety-related behavior (7), depression-like behavior (8), and avoidance behavior or receptor functions likely to be associated with differences in anxiety (9 -12). The technique of selective bidirectional breeding enhances the representation of genetic material associated with a particular trait shifting the animals' phenotype bidirectionally from the strain mean (13).Genetic approaches currently available in the mouse make this model organism particularly powerful for the functional analysis of candidate genes and in defining gene products underlying trait anxiety and possibly depression (14). For this reason we have generated and validated hyperanxious (HAB-M 1 ) and hypoanxious (LAB-M) CD1 mouse lines as model of extremes in trait anxiety and have used comparative proteomics to identify anxiety-related protein markers (2). One of the identified markers that we have reported previously, glyoxalase-I (2), showed expression level differences between HAB-M and LAB-M animals and recently has attracted increasing attention for its role in psychopathogenic mechanisms (15). The other pro...

: The identification of disease markers in tissues and body fluids requires an extensive and thorough analysis of its protein constituents. In our efforts to identify biomarkers for affective and neurological disorders we are pursuing several different strategies. On one hand we are using animal models that represent defined phenotypes charactersistic for the respective disorder in humans. In addition, we are analyzing human specimens from carefully phenotyped patient groups. Several fractions representing different protein classes from human cerebrospinal fluid obtained by lumbar puncture are used for this purpose. Our biomarker identification efforts range from classical proteomics approaches such as two dimensional gel electrophoresis and mass spectrometry to phage display screens with cerebrospinal fluid antibodies.

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