Over the last decade, a number of hydrogels attracted great attention in the area of brain tissue engineering. The hydrogels are composed of hydrophilic polymers forming 3D network in water. Their function is promoting structural and functional restoration of damaged brain tissues by providing mechanical support and navigating cell fate. This paper reports on the neurocompatibility of chitosan-g-oligo(L,L-lactide) copolymer hydrogel with primary rat cortical neuron culture. The hydrogel was produced by a molding technique on the base of photocurable composition consisting of chitosan-g-oligo(L,L-lactide) copolymer, poly(ethylene glycol) diacrylate and photosensitizer Irgacure 2959. The influence of the hydrogel on cell viability, phenotype and calcium homeostasis, mitochondrial potential and oxygen consumption rate in glutamate excitotoxicity was analyzed using primary neuron cultures obtained from a neonatal rat cortex. This study revealed that the hydrogel is non-cytotoxic. Dissociated neonatal rat cortical cells were actively attaching to the hydrogel surface and exhibited the phenotype, calcium homeostasis and mitochondrial function in both standard conditions and glutamate excitotoxicity (100 μM) similar to the control cells cultured without the hydrogel. To conclude, in this study we assessed the feasibility of the application of chitosan-g-oligo(L,L-lactide) copolymer hydrogel for tissue engineering therapy of brain injury in an in vitro model. The results support that the hydrogel is able to sustain realization of the functional metabolic activity of neonatal rat cortical cells in response to glutamate excitotoxicity.