Using an immunohistochemical technique, we have studied the distribution of 3-OH-anthranilic acid (3-HAA) in the rat brain. Our study was carried out in control animals and in rats in which a stroke model (single transient middle cerebral artery occlusion) was performed. A monoclonal antibody directed against 3-HAA was also developed. 3-HAA was exclusively observed in the infarcted regions (ipsilateral striatum/cerebral cortex), 2, 5 and 21 days after the induction of stroke. In control rats and in the contralateral side of the stroke animals, no immunoreactivity for 3-HAA was visualized. Under pathological conditions (from early phases of stroke), we reported for the first time the presence of 3-HAA in the mammalian brain. By double immunohistochemistry, the coexistence of 3-HAA and GFAP was observed in astrocytes. The distribution of 3-HAA matched perfectly with the infarcted regions. Our findings suggest that, in stroke, 3-HAA could be involved in the tissue damage observed in the infarcted regions, since it is well known that 3-HAA exerts cytotoxic effects.

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In a single transient middle cerebral artery occlusion model of stroke and using immunohistochemical techniques, the effects of a new therapeutic approach named Gemst (a member of the Poly-LLysine innovative therapies) have been studied in the rat brain. The expression of inflammatory (CD45, CD11b), oxidative (NO-tryptophan, NO2-tyrosine) and indoleamine 2, 3-dioxygenase pathway (kynurenic acid, 3-hydroxy anthranilic acid) markers has been evaluated in early and late phases of stroke. For this purpose, we have developed eight highly specific monoclonal antibodies directed against some of these markers. In the early phase (3 and 5 days of the stroke, we observed no effect of Gemst treatment (7.5 mg/day, subcutaneously for 3, 5 days). In the late phase (21 days) of stroke and exclusively in the ipsilateral side of non-treated animals an overexpression of kynurenic acid, 3- hydroxy anthranilic acid, CD45, CD11b, GFAP and ionized calcium-binding adapter molecule 1 (IBA-1) was found. In treated animals, the overexpression of the four former markers was completely abolished whereas the overexpression of the two latter ones was decreased down to normal levels. Gemst reversed the pathological conditions of stroke to normal situations. Gemst exerts a multifunctional action: down-regulates the indoleamine 2, 3-dioxygenase pathway and abolishes brain infiltration, microglial activation and gliosis. Moreover, Gemst has no effect on the expression of doublecortin, a protein involved in neuronal migration. Gemst could be a new drug for the treatment of stroke since it reverses the pathological findings of stroke and normalizes brain tissue conditions following the ischemic insult.

A highly specific monoclonal antibody directed against nitric oxide-tryptophan (NO-W) with good affinity (10-9 M) and specificity was developed. In the rat brain, using an indirect immunoperoxidase technique, cell bodies containing NO-W were exclusively found in the intermediate and dorsal parts of the lateral septal nucleus. No immunoreactive fibres were found in the rat brain. This work reports the first visualization and the morphological characteristics of cell bodies containing NO-W in the mammalian brain. The restricted distribution of NO-W in the rat brain suggests that this molecule could be involved in specific physiological mechanisms.

Introduction. The available immunohistochemical techniques have documented restricted distribution of vitamins in the mammalian brain. The aim of the study was to develop a highly specific antiserum directed against pantothenic acid to explore the presence of this vitamin in the mammalian brain. Material and methods. According to ELISA tests, the anti-pantothenic acid antiserum used showed a good affinity (10 -8 M) and specificity. The antiserum was raised in rabbits. Using an indirect immunoperoxidase technique, the mapping of pantothenic acid-immunoreactive structures was carried out in the rat brain. Results. Pantothenic acid-immunoreactive perikarya were exclusively found in the intermediate part of the lateral septal nucleus. These cells were generally small, round, fusiform or pyramidal and showed 2-3 long (50-100 µm) immunoreactive dendrites. Any immunoreactive axons containing pantothenic acid were detected. Conclusions. The very restricted anatomical distribution of the pantothenic acid suggests that this vitamin could be involved in some specific neurophysiological mechanisms.