Chemogenetic enhancement of functional recovery after a sciatic nerve injury

Abstract

Designer receptors exclusively activated by designer drugs (DREADDs) are chemogenetic tools used to modulate neuronal excitability. We hypothesized that activation of excitatory (Gq) DREADD by its designer ligand, clozapine-N-oxide (CNO), would increase the excitability of neurons whose axons have been transected following peripheral nerve injury, and that this increase will lead to an enhanced functional recovery. The lateral gastrocnemius (LG) muscle of adult female Lewis rats was injected unilaterally with AAV9- hsyn- hM3Dq-mCherry (7.6×109 viral genomes/μl) to transduce Gq-DREADD expression in LG neurons. The contralateral LG muscle served as an uninjected control. No significant changes in either spontaneous EMG activity or electrically evoked direct muscle (M) responses were found in either muscle after injection of CNO (1mg/kg, i.p.). The amplitude of monosynaptic H-reflexes in LG was increased after CNO treatment exclusively in muscles previously injected with virus, suggesting that Gq-DREADD activation increased neuronal excitability. After bilateral sciatic nerve transection and repair, additional rats were treated similarly with CNO for up to three days after injury. Electrophysiological data were collected at 2, 4 and 6 weeks after injury. Evoked EMG responses were observed as early as 2 weeks after injury only in Gq-DREADD expressing virus injected LG muscle. By 4 weeks after injury, both M-response and H-reflex amplitudes were significantly greater in muscles previously injected with viral vector than contralateral, uninjected muscles. Increases in the excitability of injured neurons produced by this novel use of Gq-DREADD were sufficient to promote an enhancement in functional recovery after a sciatic nerve injury.

Optogenetically enhanced axon regeneration: motor versus sensory neuron‐specific stimulation

“…Such a change in excitability might occur whether or not the motoneurons were actually caused to fire by afferent stimulation. Indeed, in recent evidence from our laboratory, we conclude that subthreshold increases in motoneuron excitability may be sufficient to enhance motor axon regeneration (Jaiswal & English, ). In the current experiments, selective activation of sensory neurons via optical stimulation clearly enhanced the regeneration of sensory axons but did not affect the regeneration of motor axons (neither increased nor decreased).…”

Chemogenetic Enhancement of Axon Regeneration Following Peripheral Nerve Injury in the SLICK-A Mouse

“…The simplest explanation for this chemogenetic enhancement of regeneration is that activation of the Gq DREADD using CNO resulted in an increase in the excitation of the injured neurons, leading to enhanced elongation of regenerating axons—a form of activity-dependent enhancement. In a previous study, we found that background EMG activity was not increased by CNO treatment of rats induced to express hM3Dq in motoneurons [ 14 ], leading us to conclude that activation of these modified muscarinic acetylcholine receptors produced an excitation of these neurons that was sub-threshold. Alternatively, constitutive expression of inhibitory DREADDs has been shown to induce CNO independent functional changes in sensory neurons [ 27 ].…”

“…Binding of CNO, or its major active metabolite, n-desmethylclozapine [ 9 , 10 , 11 ], to an excitatory (hM3Dq) DREADD results in activation of a Gq pathway [ 12 ] and neuronal excitation [ 13 ]. In an earlier report [ 14 ], we showed that significant enhancement of axon regeneration was found with CNO treatment in rats induced to express hM3Dq in motoneurons by retrograde adeno-associated viral transduction. Because the CNO treatments did not increase spontaneous electromyographic (EMG) activity, we concluded that DREADD activation must have produced a sub-threshold excitation in the transduced motoneurons, and that this level of excitation was sufficient to promote axon regeneration.…”

Activation of endogenous arginine vasopressin neurons inhibit food intake: by using a novel transgenic rat line with DREADDs system

“…LLC., Tokyo, Japan) was dissolved in saline (Otsuka Pharmaceutical Co. LTD., Tokyo, Japan). Dosage of CNO was determined as described previously 28,29 . At 90 min after i.p.…”