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An in vitro adult mouse muscle-nerve preparation for studying the firing properties of muscle afferents.

ABSTRACT: Muscle sensory neurons innervating muscle spindles and Golgi tendon organs encode length and force changes essential to proprioception. Additional afferent fibers monitor other characteristics of the muscle environment, including metabolite buildup, temperature, and nociceptive stimuli. Overall, abnormal activation of sensory neurons can lead to movement disorders or chronic pain syndromes. We describe the isolation of the extensor digitorum longus (EDL) muscle and nerve for in vitro study of stretch-evoked afferent responses in the adult mouse. Sensory activity is recorded from the nerve with a suction electrode and individual afferents can be analyzed using spike sorting software. In vitro preparations allow for well controlled studies on sensory afferents without the potential confounds of anesthesia or altered muscle perfusion. Here we describe a protocol to identify and test the response of muscle spindle afferents to stretch. Importantly, this preparation also supports the study of other subtypes of muscle afferents, response properties following drug application and the incorporation of powerful genetic approaches and disease models in mice.

SUBMITTER: Franco JA

PROVIDER: S-EPMC4618553 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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An in vitro adult mouse muscle-nerve preparation for studying the firing properties of muscle afferents.

Franco Joy A JA Kloefkorn Heidi E HE Hochman Shawn S Wilkinson Katherine A KA

Journal of visualized experiments : JoVE 20140924 91

Muscle sensory neurons innervating muscle spindles and Golgi tendon organs encode length and force changes essential to proprioception. Additional afferent fibers monitor other characteristics of the muscle environment, including metabolite buildup, temperature, and nociceptive stimuli. Overall, abnormal activation of sensory neurons can lead to movement disorders or chronic pain syndromes. We describe the isolation of the extensor digitorum longus (EDL) muscle and nerve for in vitro study of st ...[more]

PMID: 25285602

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Project description:Spinal afferent neurons play a major role in detecting noxious and innocuous stimuli from visceral organs, such as the gastrointestinal tract. However, all our understanding about spinal afferents has been obtained from recordings of spinal afferent axons, or cell bodies that lie outside the gut wall, or peripheral organ they innervate. No recordings have been made directly from spinal afferent nerve endings, which is where sensory transduction occurs. We developed a preparation whereby recordings could be made from rectal afferent nerve endings in the colon, to characterize mechanisms underlying sensory transduction. Dorsal root ganglia (L6-S2) were removed from mice, whilst retaining neural continuity with the colon. Fluo-4-AM was used to record from rectal afferent nerve endings in myenteric ganglia and circular muscle at 36°C. In slack (unstretched) preparations of colon, no calcium transients were recorded from spinal afferent endings. However, in response to a maintained increase in circumferential diameter, a maintained discharge of calcium transients occurred simultaneously in multiple discrete varicosities along single axons of rectal afferents in myenteric ganglia and circular muscle. Stretch-activated calcium transients were resistant to hexamethonium and nifedipine, but were abolished by tetrodotoxin, CPA, BAPTA-AM, cobalt, gadolinium, or replacement of extracellular Na(+) with NMDG. In summary, we present a novel preparation in which stretch-activated firing of spinal afferent nerve endings can be recorded, using calcium imaging. We show that circumferential stretch of the colon activates a maintained discharge of calcium transients simultaneously in varicosities along single rectal afferent endings in myenteric ganglia and circular muscle. Non-selective cation channels, TTX-sensitive Na(+) channels and both extracellular calcium influx and intracellular Ca(2+) stores are required for stretch-activated calcium transients in rectal afferent endings.

Dataset Information

An in vitro adult mouse muscle-nerve preparation for studying the firing properties of muscle afferents.

Publications

An in vitro adult mouse muscle-nerve preparation for studying the firing properties of muscle afferents.

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