Effect Of Acute Stress On Abscisic Acid In Rat Brain

Abscisic Acid (ABA) is an important universal phytohormone which regulates plants seeds dormancy, development of seedling and response to stresses including pathogens, drought, cold and salty.It was discovered recently that not only in plants, fungi, cyanobacteria and algae but also ABA exists and functions in a wide range of lower animals, such as sponges, as well as mammals including various human tissues and cells. ABA can also be produced and released from these animal cells and as a regulator on stress response, tissue regeneration, inflammation and glucose metabolism. The signal pathway in animal is remarkably similar to that in plants which ABA first binds to the ABA-binding protein located in the plasma membrane and activates intracellular ADP-ribose cyclase which would promote the production of the cyclic ADP-ribose resulting in increases of intracellular Ca²⁺ concentrations and then triggers many other correlative signal pathways. These results imply that ABA may act as an endogenous regulator in animals and may serve as a universal signal passenger in plants and animals.In order to investigate the concentration and distribution of ABA in mammalian brain, we successfully build up the method to extract and determine ABA through rat brain tissue. Using this method, we find that ABA concentration differs in different brain area including prefrontal cortex, left or right hippocampus and hypothalamus with the hypothalamus the highest ABA concentration at 18.1 1.9ng/g fresh tissue. We then study the effect of forced swim and immobilized stress on ABA level in rat brain. The results show in the immobilized 1 hour group, ABA level in prefrontal cortex and bilateral hippocampus increases twice compare to control group while decreases greatly in hypothalamus. Moreover, ABA fluctuates toward different direction between hypothalamus and other brain areas. Corresponding with other report, corticosterone in plasma increases after acute stress. Results from Q-PCR indicate the gene expression of CRF and c-fos in hypothalamus significantly up regulated in immobilized 1 hour rats. Pearson Correlation analysis shows positive correlation between changes of c-fos expression and corticosterone while negative correlation between changes of c-fos expression and ABA.We obtain three high sequence homology human proteins PPM1B, ATF6 and RNC3 when comparing by BLAST with important signal proteins ABI1, ABI3 and ABRE/ABF from A. thaliana with Positive Identities 49%,39%,60% respectively. These putative human ABA relative proteins are significantly down regulated in ABA treated BE(2)-C cells. It provides evidence that the three putative human ABA relative proteins may interact with cellular ABA signal pathway in animals.Finally ABA immunocytochemistry experiment has been done in order to discover the localization of ABA in animal brain and cells. The immunofluorescence results demonstrate that ABA is located in macrophage cellular membrane and plasma but not nucleus which is in agreement with the ABA's celluar function and the early results reported in maize roots.