Cell surface area regulation in neurons in hippocampal slice cultures is resistant to oxygen-glucose deprivation

Natalya Shulyakova^1,2, Jamie Fong², Diana Diec², Adrian Nahirny^1,2, Linda R Mills^1,2
1Department of Physiology, University of Toronto, Toronto, ON, Canada, M5T 2S8; ²Toronto Western Hospital Research Institute, University Health Network, 11-430, 399 Bathurst St, Toronto, ON, Canada, M5T 2S8

Background: Neurons swell in response to a variety of insults. The capacity to recover, ie, to shrink, is critical for neuronal function and survival. Studies on dissociated neurons have shown that during swelling and shrinking, neurons reorganize their plasma membrane; as neurons swell, in response to hypo-osmotic media, the bilayer area increases. Upon restoration of normo-osmotic media, neurons shrink, forming transient invaginations of the plasma membrane known as vacuole-like dilations (VLDs), to accommodate the decrease in the bilayer.
Methods: Here we used confocal microscopy to monitor neuronal swelling and shrinking in the three-dimensional (3D) environment of post-natal rat hippocampal slice cultures. To label neurons, we used biolistic transfection, to introduce enhanced green fluorescent protein (eGFP) targeted to the cytoplasm; and a membrane targeted GFP (lckGFP), targeted to the plasma membrane.
Results: Neurons in slice cultures swelled and shrank in response to hypo-osmotic to normo-osmotic media changes. Oxygen-glucose deprivation (OGD) caused sustained neuronal swelling; after reperfusion, some neurons recovered but in others, VLD recovery was stalled. OGD did not impair neuronal capacity to recover from a subsequent osmotic challenge.
Conclusion: These results suggest cell surface area regulation (SAR) is an intrinsic property of neurons, and that neuronal capacity for SAR may play an important role in the brain’s response to ischemic insults.

Keywords: neurons, swelling, ischemia, cell surface area, hippocampal slice culture