Ammonia-induced brain swelling and neurotoxicity in an organotypic slice model

Acute liver failure (ALF) produces cerebral dysfunction and edema, mediated in part by elevated ammonia concentrations, often leading to coma and death. The pathophysiology of cerebral edema in ALF is incompletely understood. In vitro models of the cerebral effects of ALF have predominately consisted of dissociated astrocyte cultures or acute brain slices. We describe a stable long-term culture model incorporating both neural and glial elements in a three-dimensional tissue structure offering significant advantages to the study of astrocytic-neuronal interactions in the pathophysiology of cerebral edema and dysfunction in ALF.

We utilized chronic organotypic slice cultures from mouse forebrain, applying ammonium acetate in iso-osmolar fashion for 72 hours. Imaging of slice thickness to assess for tissue swelling was accomplished in living slices with optical coherence tomography, and confocal microscopy of fluorescence immunochemical and histochemical staining served to assess astrocyte and neuronal numbers, morphology, and volume in the fixed brain slices.

Ammonia exposure at 1-10 mM produced swelling of immunochemically identified astrocytes, and at 10 mM resulted in macroscopic tissue swelling, with slice thickness increasing by about 30%. Astrocytes were unchanged in number. In contrast, 10 mM ammonia treatment severely disrupted neuronal morphology and reduced neuronal survival at 72 hours by one-half.

Elevated ammonia produces astrocytic swelling, tissue swelling, and neuronal toxicity in cerebral tissues. Ammonia-treated organotypic brain slice cultures provide an In vitro model of cerebral effects of conditions relevant to ALF, applicable to pathophysiological investigations.