**Abstract**

The H2S-producing systems were studied in trout telencephalon, tectum, and cerebellum at 1 week after eye injury. The results of ELISA analysis have shown a 1.7-fold increase in the CBS expression at 1 week post-injury, as compared to the intact trout. In the ventricular and subventricular regions of trout telencephalon, CBS+ cells, as well as neuroepithelial and glial types, were detected. As a result of injury, the number of CBS+ neuroepithelial cells in the pallial and subpallial periventricular regions of the telencephalon increases. In the tectum, a traumatic damage leads to an increase in the CBS expression in radial glia with a simultaneous decrease in the number of CBS immunopositive neuroepithelial cells detected in intact animals. In the cerebellum, we revealed neuroglial interrelations, in which H2S is probably released from the astrocyte-like cells with subsequent activation of the neuronal NMDA receptors. The organization of the H2S-producing cell complexes suggests that the amount of glutamate produced in the trout cerebellum and its reuptake is controlled with the involvement of astrocyte-like cells, reducing its excitotoxicity. We believe that the increase in the number of H2S-producing cells constitutes a response to oxidative stress, and the overproduction of H2S neutralizes the reactive oxygen species.

**Keywords:** hydrogen sulfide, traumatic eye injury, oxidative stress, radial glia, excitotoxicity, reparative neurogenesis, adult neuronal stem cells, neuroepithelial cells, astrocyte-like cells, teleost fishes, CBS expression

## **1. Introduction**

Hydrogen sulfide (H2S) was initially considered as a gasotransmitter with antioxidant properties [1]. To date, the vasodilating, neuromodulating, and anti-inflammatory properties of H2S have also been identified [2, 3]. In studies of the cardiovascular system, H2S was assumed to act as a protective factor [4]; nevertheless, the effects of H2S in the central nervous system (CNS) during stress or injury remain poorly understood. The involvement of H2S, as well as other gaseous intermediaries such as NO, CO, and H2, in the traumatic brain injury is now intensively investigated [5], but this question has not been completely clarified thus far. H2S, like nitric oxide (NO), is known to mediate posttranslational modification of proteins by adding additional sulfur to reactive cysteine residues. This modification, referred to as S-sulfhydration, is required to activate or inactivate many classes of proteins, including the ion channels, such as the ATP-dependent potassium channels, TRPV3, TRPV6, TRPM [6], enzymes, and the transcription factors NF-kB and Nrf2 [7]. Modulation of ion channels, as well as the inflammatory and the antioxidant transcription factors, using H2S after traumatic brain injury, can play a significant role in reducing edema and inflammation [8].

Recently, the involvement of H2S in cerebral ischemia, traumatic brain injury (TBI), and decrease in reactive oxygen species in the H2S-dependent mechanisms has been studied using different models [8–10]. The use of monoclonal antibodies against cystathionine β-synthase (CBS) in immunohistochemical (IHC) detection of the H2S-producing complexes in the brain of juvenile trout showed an increase in hydrogen sulfide production in different parts of the brain and CBS induction in the radial glia cells after the damage of the optic nerve [11]. It was shown that the toxic and/or neuroprotective effects of hydrogen sulfide depended on concentration: lower concentrations play a physiological role, while very high concentrations cause cell death [12, 13]. Although hydrogen sulfide is considered a gasotransmitter, there is uncertainty about the total concentration of this volatile gas or highly active anionic particles (SH-) in both plasma and central nervous system tissues [14].

The progress in studies of the hydrogen sulfide biology has led to a conclusion that polysulfides are more significant sources of intermediate sulfhydration of proteins than H2S [3]. The H2S reactions with many signal mediators, transcription factors, and channel proteins in neurons and glial cells are known both in vivo and in vitro [7, 10]. However, still little is known about interaction of the H2S intercellular communication and its consequences in the case of a traumatic cerebral injury. Such information is necessary to determine the cytoprotective or cytotoxic effects of H2S in the brain injury and/or cerebral ischemia.

The study of biology of the neural stem cells, based on animal models, is becoming increasingly important, since the processes of constitutive neurogenesis occur in many areas of the animal brain [15], providing a high reparative potential of CNS. One of such models is fish, which is characterized by a high rate of reparative processes [16]. The results of preliminary studies showed an increase in proliferative activity of cells of the trout brain after damage to optic nerve [17]. To further characterize the cellular response in the trout brain after eye injury, the hydrogen sulfide-producing enzyme, cystathionine β-synthase (CBS), was analyzed using western immunoblotting, enzyme-linked immunosorbent assay (ELISA), and immunohistochemical labeling of CBS in various sections of the trout brain at 1 week after the traumatic eye injury.
