The S1PR2‐CCL2‐BDNF‐TrkB pathway mediates neuroinflammation and motor incoordination in hyperammonaemia

Abstract

Aims: Chronic hyperammonaemia and inflammation synergistically induce neurological impairment, including motor incoordination, in hepatic encephalopathy. Hyperammonaemic rats show neuroinflammation in the cerebellum which enhances GABAergic neurotransmission leading to motor incoordination. We aimed to identify underlying mechanisms. The aims were (1) to assess if S1PR2 is involved in microglial and astrocytic activation in the cerebellum of hyperammonaemic rats; (2) to identify pathways by which enhanced S1PR2 activation induces neuroinflammation and alters neurotransmission; (3) to assess if blocking S1PR2 reduces neuroinflammation and restores motor coordination in hyperammonaemic rats.Methods: We performed ex vivo studies in cerebellar slices from control or hyperammonaemic rats to identify pathways by which neuroinflammation enhances GABAergic neurotransmission in hyperammonaemia. Neuroinflammation and neurotransmission were assessed by immunochemistry/immunofluorescence and western blot. S1PR2 was blocked by intracerebral treatment with JTE-013 using osmotic minipumps. Motor coordination was assessed by beam walking.Results: Chronic hyperammonaemia enhances S1PR2 activation in the cerebellum by increasing its membrane expression. This increases CCL2, especially in Purkinje neurons.CCL2 activates CCR2 in microglia, leading to microglial activation, increased P2X4 membrane expression and BDNF in microglia. BDNF enhances TrkB activation in neurons, increasing KCC2 membrane expression. This enhances GABAergic neurotransmission, leading to motor incoordination in hyperammonaemic rats. Blocking S1PR2 in hyperammonaemic rats by intracerebral administration of JTE-013 normalises the S1PR2-CCL2-CCR2-BDNF-TrkB-KCC2 pathway, reduces glial activation and restores motor coordination in hyperammonaemic rats.Conclusions: Enhanced S1PR2-CCL2-BDNF-TrkB pathway activation mediates neuroinflammation and incoordination in hyperammonaemia. The data raise a promising therapy for patients with hepatic encephalopathy using compounds targeting this pathway.

Extracellular Vesicles From Hyperammonemic Rats Induce Neuroinflammation in Cerebellum of Normal Rats: Role of Increased TNFα Content

“…The increased content of CCL2 induced by HA-EVs in cerebellar slices from control rats is associated with increased membrane expression and activation of its receptor CCR2 which increases BDNF in activated microglia ( Figure 7 ), in agreement with the effects reported in cerebellum of hyperammonemic rats by Arenas et al ( 22 ). The increased levels of BDNF result in increased content, membrane expression and activation of its receptor TrkB ( Figure 7 ).…”

“…We have recently shown that microglia activation in cerebellum of hyperammonemic rats is mediated by enhanced membrane expression and activation of TNFR1, which increases CCL2 production in Purkinje neurons which in turn increases activation and CCR2 membrane expression, leading to increased BDNF in microglia. BDNF released from activated microglia increases activation and membrane expression of TrkB which, in turn, enhances membrane expression of the chloride co-transporter KCC2, increasing the chloride gradient and GABAergic neurotransmission ( 22 , see Figure 7 ).…”

“…In hyperammonemia and hepatic encephalopathy peripheral inflammation induces neuroinflammation, which is prevented by preventing the appearance of peripheral inflammation ( 2 , 26 and 28 ). This neuroinflammation enhances GABAergic neurotransmission in cerebellum by two main mechanisms: increasing the TNFR1-CCL2-BDNF-TrkB-KCC2 pathway ( 22 ) and the TNFR1-NF-kB-glutaminase-GAT3 pathway ( 3 and 20 ). We show here that HA-EVs also increase these pathways, which would enhance GABAergic neurotransmission and induce motor incoordination as reported by Izquierdo-Altarejos et al ( 23 ).…”

“…This induces two main effects which will contribute to enhance G A B A e r g i c n e u r o t r a n s m i s s i o n a n d i n d u c e m o t o r incoordination: the increase in CCL2 and the nuclear translocation of the transcription factor NF-kB. The increase in CCL2 triggers microglia activation in cerebellum of hyperammonemic rats, which is prevented by blocking its receptor CCR2 (22). We show now that HA-EVs also increases CCL2 content in cerebellum of control rats and this is associated with microglia activation (Figure 7), thus reproducing the effects found in hyperammonemia.…”

“…Increased activation of TNFR1 also increases the levels of CCL2 in cerebellum, which is responsible for activation of microglia, which in turn increases BDNF levels. Increased BDNF levels enhance activation of TrkB in Purkinje neurons leading to increased membrane expression of KCC2, which enhances the transmembrane chloride gradient, further enhancing GABAergic neurotransmission ( 22 ).…”