Analysis of endocannabinoid receptors and enzymes in the post-mortem motor cortex and spinal cord of amyotrophic lateral sclerosis patients.

Objective: We have investigated the endocannabinoid system in the motor cortex of motor neuron disease (MND) patients.
Methods: Post-mortem samples from MND patients and controls were used for immunostaining and/or Western blotting analysis of endocannabinoid elements.
Results: We did not find any evidence of neuronal losses in the motor cortex of MND patients, but elevations in glial markers Iba-1 and GFAP were evident. We found no changes in FAAH and MAGL enzymes and in the CB ₁ receptor, which correlated with the lack of cortical neuron death. By contrast, the Western blotting analysis of CB ₂ receptors proved an increase in the motor cortex corroborated by immunostaining, correlating with the elevated gliosis in these patients. Double-labeling analyses revealed that this elevated CB ₂ receptor immunostaining was located in GFAP-labelled astroglial cells. However, we also found CB ₂ receptor labeling in cortical neurons confirmed with double immunofluorescence with the neuronal marker MAP-2. This was also found in the spinal cord, using double-labeling with the spinal motor neuron marker choline-acetyl transferase. This happened in both patients and controls, despite these neurons experienced an important degeneration in patients reflected in reduced Nissl staining, TDP-43 immunostaining and CB ₁ receptor levels measured by Western blotting.
Conclusion: We have confirmed that CB ₂ receptors are elevated in the motor cortex of MND patients associated with the reactive gliosis. This phenomenon is previous to neuronal losses. We also found CB ₂ receptors in cortical and spinal motor neurons. These observations support that targeting this receptor may serve for developing neuroprotective therapies in MNDs.