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1. Distribution of c-Fos positive nuclei in: ventral tegmental area, substantia nigra, periaqueductal gray. 2. Appearance of calbindin-D28k, calretinin and parvalbumin in midbrain neurons that are engaged in the stress response. 3. Changes of c-Fos and calcium-binding proteins expression during maturation. The material consisted of Wistar rats of age between 0 and 90 days. The OF exposure was applied throughout 10 min and 90 min before the death of the animals. The brain sections were double stained using the antibodies against c-Fos, CB, CR or PV. Our results showed that in all studied nuclei age-related increase of c-Fos expression (without changing of its distribution properties) was found. PV didn't show any co-localization with c-Fos in neurons of studied regions at any ages, however some PV-immunoreactive (PV-ir) basket-like structures around c-Fos-immunoreactive (c-Fos-ir) neurons were observed. In the youngest group of rats c-Fos-ir cells and cells immunoreactive for CB and CR constituted separate neuronal populations. During maturation increases in the level of their co-localization with c-Fos was observed. We may conclude that in adult rat midbrain structures CB-immunoreactive (CB-ir) and CR-immunoreactive (CR-ir) cells (probably projection neurons) are mainly activated in the stress response following OF exposure. In the contrary PV-ir cells has only an indirect (modulatory) influence upon the c-Fos-ir cells.
In our study we used c-Fos protein (as a marker of cellular activity) to identify whether cells containing parvalbumin (PV) in the piriform cortex (PC) are engaged in the response to stress stimulation and to discover how this expression changes during maturation. The material consisted of Wistar rats of postnatal (P) ages between 0 and 120 days divided into 9 groups: P0, P4, P7, P10, P14, P21, P30, P90, P120. Each group consisted of 5 experimental and 3 control animals. Rats of the experimental groups were exposed to the “open field test” throughout 10 minutes. The control animals were kept in a home cage. Our results showed that c-Fos activity in the open field test was observed in layers II and III of PC after birth. It then increased and stabilised on P30. In the second week of life PV-positive cells were also observed in those layers. These achieved maturity in the 4th week of life. After this time basket-like structures appeared but the level of PV/c-Fos co-localisation was low. Only small differences were observed between the anterior and posterior parts of PC. In the anterior part a higher number of PV-positive neurons, neuropil threads, and basket-like structures and a larger degree of PV/c-Fos co-localisation were observed. Our results suggested that during maturation PV cells are not directly activated in response to stress stimuli but PV neurons via their numerous endings influence the activation of c-Fos-positive cells predominantly in the anterior part of PC.
Our intention in the present study was to ascertain whether NO-producing cells in the basolateral complex (BLC) and paracapsular intercalated nerve cell groups (Ip) of the amygdala are activated in the open field (OF) test. The material consisted of 8 adult rat brains. The OF test was applied throughout 10 min and 90 min before the death of the animals. The brain sections were double stained using the antibodies against c-Fos (marker of neuronal activation) and against nitric oxide synthase (NOS — marker of NO-producing cells). The neurons containing NOS and those revealing c-Fos activity constituted distinct populations within both the BLC and Ip but NOS-immunoreactive fibres often surrounded the c-Fos-immunoreactive neurons. Our results suggest that (1) neurons of the basolateral complex of the amygdala and paracapsular intercalated islands are involved but probably not crucial for the open field stress response and (2) NOS-immunoreactive cells in the BLC and Ip are not activated after OF exposure.
In the present study we wanted to check whether the expression of the c-Fos protein (the marker of cellular activity) appears in cells containing calcium-binding proteins (CaBPs) in animals exposed to the open field test. Eight adult Wistar rats were examined. In the first step the open field test was applied throughout 10 minutes. After perfusional fixation brains were frozen and cut on the cryostat in the coronal plane and stained with the standard immunohistochemical method. Sections were double stained for c-Fos and CaBPs: parvalbumin (PV), calbindin (CB), calretinin (CR). c-Fos positive cells were localized predominantly in layers II and III of the piriform cortex (PC). The double labeling study showed that neurons containing CaBPs are rarely c-Fos-immunoreactive. Often PV-positive and CB-positive fibers surround c-Fos-positive neurons in layers II and III in a form of a basket. It seems that cells containing CaBPs are not directly involved in the response to aversive stimuli but cells containing those calcium-binding proteins might influence directly c-Fos positive neurons of PC.
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