Effects of stress on rat brain adenosine 3',5'-monophosphate in vivo.

The stress of electric foot shock or restraint increases the rate of metabolism in vivo of norepinephrine (NE), dopamine (DA), and serotonin in subsections of rat brain 1. Since these substances as well as other proposed neurotransmitters stimulate the accumulation of adenosine Y,5'-monophosphate (cAMP) in nervous tissue in vitroS,l°-12,1~,a7, it was decided to measure the effects of foot shock or restraint on the concentration of cAMP in various regions of the brain. Previously, studies of the cAMP system in vivo have been hampered by the marked increases which occur in brain cAMP levels immediately after death when animals are not frozen or are decapitated before freezing2,9, la. Quick freezing of the whole animal in liquid nitrogen reduces these changes significantly, but has generally restricted the level of analysis to the whole brain since the hard-frozen tissue is difficult to dissect 9,1a. Analyses of various regions of the rat brain have been performed using microwave fixation, but this method may produce significant tissue anoxia and hydrolysis of cAMP la. A method recently developed in this laboratory, direct liquid nitrogen freezing followed by softening of the tissue in anhydrous trichloroacetic acid (TCA) in acetone, allows both quick fixation and accurate dissection of the tissue 5. We have found marked increases in cAMP levels in the septum of rats subjected to electric foot shock or restraint; smaller increases were observed in the hippocampus and brain stem of shocked animals only. Male Sprague-Dawley rats, weighing 200-250 g, were kept in experimentalcontrol pairs in the same cage for at least 36 h prior to experimental treatment. Shocked animals received a 25 mV DC, 4 mA, 1-sec duration foot shock from a floor grid every 10 sec for 1 h I. Restrained animals were wrapped firmly in wire mesh for 2 h such that head, tail and limb movements were minimal and breathing was not restricted. Animals were killed immediately after treatment by immersion head-first into liquid nitrogen for at least 5 min. Control animals received approximately the same handling as experimentals but remained in the home cage in a separate room during the treatment period. Each control was killed as described within 10 min of its cage-