**2.3. The estimation of blood cortisol and catecholamine (CA) levels**

Blood samples for the analysis of CA estimation were collected from the jugular external vein (according to a scheme described above—**Figure 1**). Blood samples were placed in 10 mL test tubes containing reduced glutathione (0.05 mM). The plastic tubes were maintained on ice, and after the centrifugation, plasma was stored at −80°C, until the beginning of the analytical process. The detection of CA levels was performed by radioimmunoassay using REA kits (CATECHOLA, Czech Republic). The sensitivity of this method was for E, 0.37, for NE, 0.53, and for DA, 0.85 nM L−1. The intra serial error for E was of 3%, for NE 4.2%, and for DA 6.1%, whereas the error among the series was of 4.2, 7.4, and 6.6%, respectively.

Cortisol levels were detected by radioimmunoassay (RIA), according to previous experiments [18]. The mean intra and inter assay of the method was of 9.5 pg, for a sample of 10 μL (ORION DIAGNOSTICA, Espoo). Δmax concentration for each hormone was the difference between the basal concentration and the highest concentration measured.
