Chronic G_q-DREADD Activation Causes Cell Loss

Luigi Bellocchio, Andrea Ruiz-Calvo, Anna Chiarlone, Magali Cabanas, Eva Resel, et al.

(see pages 10611–10624)

Motor behaviors are coordinated by two classes of striatal medium spiny neurons (MSNs). Direct-pathway MSNs facilitate movements by disinhibiting basal ganglia targets, while indirect-pathway MSNs suppress movements by enhancing target inhibition. Activity in both MSN classes is modulated by neurotransmitters that bind to g-protein coupled receptors. For example, dopamine increases activity in direct-pathway MSNs (D1R-MSNs) via G_olf-coupled D1 receptors and decreases activity in indirect-pathway MSNs (D2R-MSNs) via G_i-coupled D2 receptors.

Both MSN classes also express Group I metabotropic glutamate receptors (mGluRs) and type 1 muscarinic acetylcholine receptors, which are coupled to G_q. Unlike G_olf and G_i, which signal through increases and decreases in cAMP, respectively, G_q signals via phospholipase C and increased intracellular calcium levels. Although excessive mGluR-dependent signaling has been proposed to contribute to MSN loss in Huntington's disease (Dowie et al. 2010 Pharmacol Ther 128:305), the intracellular signaling cascades activated in MSNs by these receptors are poorly understood. To address this, Bellocchio et al. expressed hM3Dq, a G_q-coupled designer receptor exclusively activated by designer drugs (DREADD), in mouse D1R- or D2R-MSNs.

Acute activation of hM3Dq in D1R-MSNs increased daytime activity and reduced sleep. Conversely, acute hM3Dq activation in D2R-MSNs reduced activity and increased sleep. These effects disappeared with chronic activation of G_q signaling, and in D2R-MSNs, prolonged (14 d) G_q activation caused mice to become more active and sleep less than controls—the opposite of the acute effects. These changes were associated with a decrease in MSN numbers after chronic G_q activation. In vitro experiments revealed a hM3Dq-dependent neurotoxic pathway involving activation of phospholipase C, release of calcium from internal stores, activation of the tyrosine kinase PYK2, and phosphorylation/activation of JNK kinase. Consistent with this, a JNK inhibitor prevented neuronal loss and the reversal of behavioral effects during chronic G_q activation in vivo.

These results show that chronic activation of G_q signaling can kill MSNs by activating JNK. Such cytotoxic effects should be considered in future experiments using chronic DREADD activation. Nonetheless, previous studies either found no effect of chronic G_q activation or attributed diminished effects to receptor desensitization. An intriguing possibility is that MSNs are particularly sensitive to chronic Gq signaling, which may contribute to their susceptibility to death in Huntington's disease.

Neural Precursors Improve Ischemic Outcome via VEGF

Marco Bacigaluppi, Gianluca Luigi Russo, Luca Peruzzotti-Jametti, Silvia Rossi, Stefano Sandrone, et al.

(see pages 10529–10544)

Cerebral artery obstruction leads to excessive glutamate release and excitotoxicity, causing functional impairment. Often function is partially restored by plasticity in spared circuits, however. Importantly, the extent of recovery can be increased by treatments that limit excitotoxicity and enhance plasticity. One promising treatment in this regard is neural precursor cell (NPC) transplantation. NPCs can promote recovery not only by generating new neurons, but also by secreting growth factors and neuroprotective molecules. Bacigaluppi et al. demonstrate the latter effect in a mouse model of stroke.

View larger version:

• In this window
• In a new window

• Download as PowerPoint Slide

Reconstructions of the peri-ischemic area (lesion border indicated by dashed white line) show that GLT-1 (green) expression is higher 10 d after NPC injection (bottom) than at the same time in controls (top). See Bacigaluppi et al. for details.

Three days after middle cerebral artery occlusion, NPCs were injected intravenously. NPCs migrated to the lesion area and remained there, undifferentiated, for at least 60 d. Compared to controls, NPC-treated mice had smaller lesion volumes, less degeneration in the corticospinal tract, and more sensory and motor improvement. Furthermore, the dendritic arbors of corticospinal tract neurons were longer in both ipsilesional and contralesional hemispheres and sprouting of contralesional cortical axons towards denervated targets was greater in NPC-treated mice than controls.

Arterial occlusion reduced the frequency and amplitude of spontaneous IPSCs and increased the frequency and amplitude of EPSCs in striatal medium spiny neurons (MSNs), which receive cortical input. In addition, the proportion of EPSCs carried by NMDA receptors was reduced in ipsilesional striatum. NPC treatment reduced ischemic effects on IPSC frequency, IPSC amplitude, and EPSC amplitude in ipsilesional MSNs. It also increased neuronal survival in the ipsilesional striatum and increased NMDA-receptor currents in contralateral MSNs.

The beneficial effects of NPCs likely stemmed from their secretion of vascular endothelial growth factor (VEGF), which upregulated the glutamate transporter GLT-1 in astrocytes, resulting in reduced extracellular glutamate. Blocking VEGF with a neutralizing antibody reduced GLT-1 upregulation, and a GLT-1 inhibitor both prevented the NPC-induced enhancement of NMDA-receptor currents and eliminated the beneficial effect of NPC treatment on functional recovery.

These results complement previous work showing that VEGF and GLT-1 enhance plasticity and reduce damage resulting from ischemia, and they demonstrate that NPCs can improve stroke outcomes without differentiating into neurons. Given that NPCs were not detected in the contralesional hemisphere, the enhancement of contralesional plasticity is surprising, Future work should investigate the mechanisms underlying this effect.