Risk assessment (RA) consists of rodents’ defense behaviors displayed towards aversive stimuli, and is enhanced by anxiogenic drugs and reduced by anxiolytics. Although scoring of RA has been proposed to improve the sensitivity of the elevated plus maze (EPM) test to drugs that modulate anxiety [1], the lack of methodological standardization means that RA has limited use as a reliable parameter to be analyzed alongside the conventional measures of time and entries in open and closed arms of the EPM. To try to show the importance of a deeper investigation into RA measurement for enhancing EPM sensitivity, rats were tested under the effect of four drugs: two related to the benzodiazepine-GABA neurotransmitter system; and two which act on NK-1 receptors, but have as yet no established effect on human anxiety.

Naïve male Wistar rats received intra-peritoneal injections of either the anxiolytic diazepam, the anxiogenic pentylenetetrazol, the NK-1 agonist (substance P, or Sar9Met[O_2-11]-substance P), the NK-1 antagonist (GR82334, given alone or with substance P), or a control vehicle. After a fixed amount of time, each animal was allowed to freely explore the EPM for 5 min. During this period, conventional measures of time and number of entries, as well as distance moved in the EPM zones (open and closed arms, center zone, and open arm ends), were automatically recorded by the EthoVision video-tracking tool. At the same time, RA (postures with stretched body, flat back and elongation of the head) was observed and scored with the EthoVision manual event recorder tool, in terms of absolute duration and frequency in each zone. These were then converted into percentage measures, % RA time and % RA frequency, calculated as factors of the time spent in each zone and of the total number of occurrences displayed over the whole session, respectively.

Rats that received diazepam (0.3, 1 and 3 mg/kg) displayed significantly less RA than controls, but no dose was able to significantly change conventional measures. Pentylenetetrazol (1, 3, 10 and 20 mg/kg) enhanced RA, but only the highest dose increased time spent in the closed arms. The NK-1 agonists (50 µg/kg) enhanced time and distance moved in the open arms, but each drug changed different RA parameters. Both open activity-enhancing and RA-reducing actions of substance P (considered to be anxiolytic effects) were abolished by pre-treatment with GR82334. The most sensitive RA parameters were % RA time in the open arms and in the open arm ends.

These results provide evidence that measuring the RA of rats in the EPM is advantageous to achieving an appropriate sensitivity to both typical and novel drugs. It can also be practical and reliable, especially when combined with a tracking tool that provides spatio-temporal measures.

References

1. Rodgers, R.J.; Dalvi, A. (1997). Anxiety, defense and the elevated plus-maze. Neuroscience and Biobehavioral Reviews, 21(6), 801-10.

Paper presented at Measuring Behavior 2002 , 4^th International Conference on Methods and Techniques in Behavioral Research, 27-30 August 2002, Amsterdam, The Netherlands

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