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. Author manuscript; available in PMC: 2018 Jul 1.

Published in final edited form as: Anesth Analg. 2017 Jul;125(1):241–254. doi: 10.1213/ANE.0000000000002137

Insufficient BDNF in the low density of astrocytes in the co-culture contributes to propofol-induced neuron death through BDNF/GSK3β/mitochondrial fission pathway. (A) Propofol-induced increase of mitochondrial fission contributes to neuron death in both neuron-alone culture and astrocyte-neuron co-culture with the low-density of astrocytes (ratio of astrocytes/neurons is 1:9). (A-a) The representative confocal fluorescence images of mitochondria in neuron-alone culture and 1:9 neuron-astrocyte co-culture. Following 6 h of 30 μM propofol exposure, mitochondria were labeled with the mitochondrial dye TOM20 (green) using immunofluorescence staining and imaged by confocal microscopy. The blue are cell nuclei stained with Hoechst 33342. In the control and Mdivi-1-treated neurons, the mitochondria were interconnected and tubular. Following propofol exposure, much shorter and fragmented mitochondria exhibiting punctate signals were prevalent in the cells. (A-b & c) Quantification of mitochondrial shape in the neurons by measuring both aspect ratio and form factor represent mitochondrial length and mitochondrial branching, respectively. Propofol significantly decreased the values of aspect ratio and form factor in neuronal processes of neuron-alone culture and 1:9 astrocyte-neuron co-culture, indicating enhanced mitochondrial fission while Mdivi-1 prevented propofol-induced mitochondrial fragmentation by increasing both aspect ratio and form factor. (A-d) Inhibition of mitochondrial fission by Mdivi-1 rescued propofol-induced neuron death in neuron-alone culture and 1:9 astrocyte-neuron co-culture. (B) The effect of BDNF asministration and co-cultured high-density of astrocytes on propofol-induced mitochondrial fission in neurons. (B-a & b) BDNF administration attenuated the propofol-induced mitochondrial fission by increasing both aspect ratio and form factor in neuron-alone culture and 1:9 astrocyte-neuron co-culture. The high density of astrocytes attenuated the propofol-induced mitochondrial fragmentation in neurons (ratio of astrocytes/neurons is 1:1). (C) The effect of GSK3β inhibitor CHIR99021 on propofol-induced mitochondrial fission in neuron-alone culture and 1:9 astrocyte-neuron co-culture. (C-a & b) CHIR99021 attenuated the propofol-induced neuron mitochondrial fission by increasing both aspect ratio and form factor. n=5. *P<0.05.

Insufficient BDNF in the low density of astrocytes in the co-culture contributes to propofol-induced neuron death through BDNF/GSK3β/mitochondrial fission pathway. (A) Propofol-induced increase of mitochondrial fission contributes to neuron death in both neuron-alone culture and astrocyte-neuron co-culture with the low-density of astrocytes (ratio of astrocytes/neurons is 1:9). (A-a) The representative confocal fluorescence images of mitochondria in neuron-alone culture and 1:9 neuron-astrocyte co-culture. Following 6 h of 30 μM propofol exposure, mitochondria were labeled with the mitochondrial dye TOM20 (green) using immunofluorescence staining and imaged by confocal microscopy. The blue are cell nuclei stained with Hoechst 33342. In the control and Mdivi-1-treated neurons, the mitochondria were interconnected and tubular. Following propofol exposure, much shorter and fragmented mitochondria exhibiting punctate signals were prevalent in the cells. (A-b & c) Quantification of mitochondrial shape in the neurons by measuring both aspect ratio and form factor represent mitochondrial length and mitochondrial branching, respectively. Propofol significantly decreased the values of aspect ratio and form factor in neuronal processes of neuron-alone culture and 1:9 astrocyte-neuron co-culture, indicating enhanced mitochondrial fission while Mdivi-1 prevented propofol-induced mitochondrial fragmentation by increasing both aspect ratio and form factor. (A-d) Inhibition of mitochondrial fission by Mdivi-1 rescued propofol-induced neuron death in neuron-alone culture and 1:9 astrocyte-neuron co-culture. (B) The effect of BDNF asministration and co-cultured high-density of astrocytes on propofol-induced mitochondrial fission in neurons. (B-a & b) BDNF administration attenuated the propofol-induced mitochondrial fission by increasing both aspect ratio and form factor in neuron-alone culture and 1:9 astrocyte-neuron co-culture. The high density of astrocytes attenuated the propofol-induced mitochondrial fragmentation in neurons (ratio of astrocytes/neurons is 1:1). (C) The effect of GSK3β inhibitor CHIR99021 on propofol-induced mitochondrial fission in neuron-alone culture and 1:9 astrocyte-neuron co-culture. (C-a & b) CHIR99021 attenuated the propofol-induced neuron mitochondrial fission by increasing both aspect ratio and form factor. n=5. *P<0.05.