c-Fos expression induced by electroacupuncture at the Zusanli point in rats submitted to repeated immobilization

Abstract

In laboratory animals, acupuncture needs to be performed on either anesthetized or, if unanesthetized, restrained subjects. Both procedures up-regulate c-Fos expression in several areas of the central nervous system, representing therefore a major pitfall for the assessment of c-Fos expression induced by electroacupuncture. Thus, in order to reduce the effect of acute restraint we used a protocol of repeated restraint for the assessment of the brain areas activated by electroacupuncture in adult male Wistar rats weighing 180-230 g. Repeated immobilization protocols (6 days, 1 h/day and 13 days, 2 h/ day) were used to reduce the effect of acute immobilization stress on the c-Fos expression induced by electroacupuncture at the Zusanli point (EA36S). Animals submitted to immobilization alone or to electroacupuncture (100 Hz, 2-4 V, faradic wave) in a non-point region were compared to animals submitted to electroacupuncture at EA36S (4 animals/subgroup). c-Fos expression was measured in 41 brain areas by simple counting of cells and the results are reported as number of c-Fos-immunoreactive cells/10,000 µm 2 . The protocols of repeated immobilization significantly reduced the immobilizationinduced c-Fos expression in most of the brain areas analyzed (P < 0.05). Animals of the EA36S groups had significantly higher levels of c-Fos expression in the dorsal raphe nucleus, locus coeruleus, posterior hypothalamus and central medial nucleus of the thalamus. Furthermore, the repeated immobilization protocols intensified the differences between the effects of 36S and non-point stimulation in the dorsal raphe nucleus (P < 0.05). These data suggest that high levels of stress can interact with and mask the evaluation of specific effects of acupuncture in unanesthetized animals.

Opioidergic orofacial antinociception induced by electroacupuncture at acupoint St36

“…With the aim to test this hypothesis, our group conducted an experiment that did not show the influence of stress in our experimental groups. Thus, this hypothesis was not confirmed by our experimental data, which are in accordance with other studies that used the same restraint method (results not shown) (17,18).…”

“…Rats were immobilized in a plastic cylinder that allowed access to all four limbs for the application of acupuncture needles (17,18). This apparatus permits acupuncture stimulation in awake rats.…”

“…However, less is known about its action on trigeminal pain. There are few studies on humans concerning orofacial pain and even fewer controlled laboratory and animal studies (12,13 The objectives of the present study were: a) to determine the efficacy of acupuncture antinociception in a model of orofacial nociception in rats using points recognized to be analgesic, St36 (14)(15)(16)(17)(18) and LI4 (19)(20)(21)(22), and b) to assess if this effect occurs via endogenous opioid mobilization.…”

Stress-Induced c-Fos Expression is Differentially Modulated by Dexamethasone, Diazepam and Imipramine

“…Immobilization (2 h) for 13 consecutive days was able to reduce the immobilizationinduced c-Fos expression in most brain structures, including amygdala, hypothalamus, thalamic nuclei, and cortical areas (Medeiros et al, 2003). In this sense, the wide effect of repeated stress can be related to the fact that the habituation acted simultaneously in many circuits independently of which neurotransmitters or intracellular mechanisms were involved.…”

“…Stereological methods were not employed in this study due to potential bias associated with counts generated in this manner, such as uncertainties as to the extent to which antiserum penetrates through the thickness of the tissue sections and difficulties in defining the boundaries of the several cell groups of interest. Moreover, our interest was to make only relative comparisons of the strength of Fos induction as a function of the treatment status (Li and Sawchenko, 1998;Medeiros et al, 2003).…”