No abstract

BACKGROUND AND PURPOSEThe role of inosine at the mammalian neuromuscular junction (NMJ) has not been clearly defined. Moreover, inosine was classically considered to be the inactive metabolite of adenosine. Hence, we investigated the effect of inosine on spontaneous and evoked ACh release, the mechanism underlying its modulatory action and the receptor type and signal transduction pathway involved. EXPERIMENTAL APPROACHEnd-plate potentials (EPPs) and miniature end-plate potentials (MEPPs) were recorded from the mouse phrenic-nerve diaphragm preparations using conventional intracellular electrophysiological techniques. KEY RESULTSInosine (100 μM) reduced MEPP frequency and the amplitude and quantal content of EPPs; effects inhibited by the selective A3 receptor antagonist MRS-1191. Immunohistochemical assays confirmed the presence of A3 receptors at mammalian NMJ. The voltage-gated calcium channel (VGCC) blocker Cd 2+ , the removal of extracellular Ca 2+ and the L-type and P/Q-type VGCC antagonists, nitrendipine and ω-agatoxin IVA, respectively, all prevented inosine-induced inhibition. In the absence of endogenous adenosine, inosine decreased the hypertonic response. The effects of inosine on ACh release were prevented by the Gi/o protein inhibitor N-ethylmaleimide, PKC antagonist chelerytrine and calmodulin antagonist W-7, but not by PKA antagonists, H-89 and KT-5720, or the inhibitor of CaMKII KN-62. CONCLUSION AND IMPLICATIONSOur results suggest that, at motor nerve terminals, inosine induces presynaptic inhibition of spontaneous and evoked ACh release by activating A3 receptors through a mechanism that involves L-type and P/Q-type VGCCs and the secretory machinery downstream of calcium influx. A3 receptors appear to be coupled to Gi/o protein. PKC and calmodulin may be involved in these effects of inosine. Abbreviationsαβ-MeADP, α,β-methyleneadenosine 5'-diphosphate; ω-CgTx, ω-conotoxin GVIA; ACh, acetylcholine; BgTx-R, α-bungarotoxin coupled to tetramethylrhodamine; CAMKII, calcium/calmodulin-dependent protein kinase

In this work, we built ultrasonic disc-shaped transducer for targeted neuromodulation with the addition of a solid axicon lens based on a polydimethylsiloxane (PDMS) interface. We made a numerical and experimental characterization of its acoustic field. The motor cortex of CF-1 mice was stimulated, through the skin and skull into the intact brain, with low-intensity pulsed ultrasound. Evoked muscle responses in different body segments were clearly observed, including hindlimb, forelimb, and tail. Axicon lens affixed on the face of the transducer makes possible a targeted modulation of the motor cortex by pulsed ultrasound, inducing muscle contraction in a specific body segment. In this approach, the lateral and axial spatial resolution is comparable to spherical segment ultrasound transducers, but with a shorter focal length. Thus, ultrasound axicon looks attractive to investigate the functional contributions of fine-grained spatial structures in the brain.