The isometric tetanic force of all mice TA muscles were measured with a commercial device (Grass Tech, Astro-Med Inc) as previously described63 (link). Briefly, mice were anesthetized by isoflurane and warmed by a heating lamp during the entire procedure, the tendon was exposed and attached to a force transducer (Grass FT03 Transducer, Astro-Med Inc), and the knee was immobilized by a stainless steel pin. The electrical stimulation was performed via a bipolar electrode with a Grass stimulator to the sciatic nerve. The maximum isometric tetanic force was achieved by applying single-pulse stimuli (volts = 12 V, duration = 0.2 ms, pulse rate = 100 Hz) at an optimal muscle length, which was adjusted with 0.5 mm increments. Data were acquired and recorded with the PolyVIEW16 software (Grass Tech, Astro-Med Inc.). Following the completion of the isometric force testing, the mouse was euthanized, and the entire TA muscle was carefully dissected and weighed.
Grass stimulator
Manufactured by Natus
Sourced in United States
The Grass stimulator is a specialized laboratory equipment used to generate electrical impulses for various research and experimental purposes. It is designed to provide controlled and programmable stimulation signals to study physiological responses in research settings.
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19 protocols using grass stimulator
1
Isometric Tetanic Force Measurement in Mouse TA Muscle
2
Hippocampal CA3-CA1 Circuit Stimulation
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A bipolar electrode, made of polyimide‐insulated stainless steel wire, was placed in the CA3 striatum oriens area for stimulating the CA3 circuitry. Constant current pulses with durations of 0.1 ms were generated by the Grass stimulator and delivered every 30 sec through the isolation unit as described above. CA1 field EPSPs and paired pulse facilitation were evoked by paired stimuli at a low intensity (45–70 μA) with an interpulse interval of 50 ms. CA3 population spikes and paired pulse depression (PPD) were evoked by paired stimuli at the maximal intensity (150 μA) with an interpulse interval of 250 ms. Strong stimuli were used in the latter protocol because slow IPSCs sensitive to blockade by CGP 55845 were reliably induced by single stimulation at the maximal intensity (see below). The interpulse interval of 250 ms was modified from previous studies of GABAB‐mediated depression of hippocampal glutamatergic responses (Davies et al. 1993 ; Isaacson et al. 1993 (link); Leung et al. 2008 (link)). To induce repetitive field potentials in conventional slices, we used a train of high‐frequency stimuli at the maximal intensity (80 Hz for 1 sec, current pulses of 150 μA).
3
ECC-induced Muscle Hypertrophy in Mice
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Eccentric contractions of the left tibialis anterior (TA) muscle was induced by high‐frequency electrical stimulation of the sciatic nerve as previously described with slight modifications.16 , 24 Mice were anaesthetized via isoflurane (2% in O2 with 1.5% maintenance), the left leg/hip region was shaved, and two needle electrodes were placed subcutaneously on the left leg to stimulate the sciatic nerve. Tetanic muscle contractions were generated using a Grass Stimulator (Model S88, Grass Instruments, Quincy, MA, USA) for 10 sets of six repetitions (100 Hz, 6–12 V, 1 ms duration). Ten seconds of rest was given between repetitions, and 50 s of rest was given between sets. The stimulation protocol recruits all motor units and results in net plantar flexion of the ankle.33 , 34 The dorsiflexors (TA and EDL) undergo ECCs while the plantar flexors (gastrocnemius, soleus, and plantaris) perform concentric muscle contractions. In all experiments, the left TA performed ECC while the right TA served as intra‐animal, non‐contracted control. Our laboratory and others have demonstrated repeated ECC bouts, but not concentric contractions, can induce muscle and myofiber growth in rodents.16 , 23 , 24 , 33 Thus, the TA was examined in all experiments. Mice were given an intraperitoneal injection of warm saline following the stimulation procedure and returned to cages upon complete recovery.
4
Stimulation of Crayfish Opener Muscle
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The excitatory axon innervating the opener muscle in the crayfish was stimulated in the meropodite by placing a branch of the leg nerve into a suction electrode connected to a Grass stimulator [20 (link)]. Stimulation at 40-Hz trains of 30 pulses in duration was applied to the excitatory nerve to compare the responses obtained in the three general regions of the opener muscle: distal, central, and proximal). Preparations were used immediately after dissection, and all the experiments were performed at room temperatures (19–21 °C).
5
Sciatic Nerve Denervation in Rats
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In a third aseptic surgical procedure, the skin was shaved on both hindlimbs and then opened to expose, bilaterally, the sciatic nerve and it’s tibial (TIB), common peroneal (CP) and sural nerve branches. An aseptic bipolar electrode attached to a Grass stimulator was used to deliver supramaximal stimuli at 10 Hz to the proximal sciatic nerve stump, followed by the distal stumps of the sciatic nerve and its branches to ensure that nerve regeneration had not occurred. No muscle contractions were observed in any of the rats confirming that the distal nerve stumps of the sciatic nerves were acutely denervated for 7 days and chronically denervated for 7 weeks and 17 months.
6
Muscle Hyperalgesia Induction Protocol
7
Electrical Stimulation of Vagal Nerve
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The right cervical vagus nerve was carefully dissected free from surrounding structures and cut just caudal to the nodose ganglion, then the caudal, cut end was placed on a pair of bipolar platinum hook electrodes and electrically stimulated with a Grass Stimulator (S48). The stimuli [square wave pulses (30 μA, 1 ms) at 2–30 Hz for 20 s] were delivered via an isolation unit (SIU 6). After each stimulation of the left ADN or right vagal nerve, HR and AP returned to their pre-stimulus baseline levels. Maximal HR and MAP responses to electrical stimulation of the cervical vagal nerve were measured. The data were analyzed as described for ADN stimulation. The stimulation-induced changes in HR and MAP were abolished after the vagus nerve was crushed caudal to the electrode, confirming that the responses were indeed due to vagal efferent activity.
8
Identifying Cardiac Ganglionated Plexus
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The detailed preparation procedures have been reported before.10 , 17 , 18 , 19 In brief, after the lateral thoracotomy, the GPs were identified by applying high‐frequency stimulation (HFS) from a Grass stimulator applied with a bipolar electrode probe (20‐Hz, 0.1‐ms‐duration square waves, 0.6–8.0 V). The fat pad containing the anterior right GP (ARGP) was situated between the caudal end of the sinoatrial node and RSPV–atrium junction. The inferior right GP (IRGP) was located at the junction of the inferior vena cava and both atria. The superior left GP (SLGP) was located adjacent to the LSPV–atrium junction between the LA appendage and left PA. The inferior left GP (ILGP) was located on the caudal side of the LIPV–atrium junction. A bradycardic response, which showed a progressive slowing of the sinus rate by 50% or the development of second‐ or third‐degree atrioventricular block resulting from incremental voltage levels applied to the fat pad, was used as a surrogate marker for GP stimulation.
9
Electrical Pulp Stimulation in Stereotaxic Instrument
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After placing the animal’s head in a stereotaxic instrument, the tips of both lower incisors were cut off with a dental separator, and stainless-steel wire electrodes were inserted into the pulp and fixed with dental cement. The bipolar pulp stimulation was delivered 6 times per minute, with a train of four electrical impulses of 200 Hz frequency, for a duration of 3 ms per single impulse with 2 ms intervals and a 4–5 V amplitude, using a programmed stimulator. Trains of 4 impulses were delivered to the pulp at 10 s intervals using a Grass stimulator coupled with a gate generator.
10
Evaluating Cardiomyocyte Contractility and Calcium Dynamics
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Cell culture media was replaced with 37°C Tyrode’s salt solution (supplemented with 0.1% DMSO with or without 50 μg/mL DEHP or 100 μM Wy-14,643), and spontaneous beating rate recordings were collected using a Zeiss LSM 510 confocal imaging system (488 nm excitation/505-550 nm emission filters). Cells were then equilibrated at room temperature for 20 min [19 (link)], and pace-induced calcium transient recordings were measured. In the latter, the cell network was paced using a stimulation electrode (Harvard Apparatus, Holliston MA) to which monophasic 5 msec pacing pulses were applied (4V minimum threshold, Grass Stimulator). In a second set of studies, cardiomyocytes were exposed to 20 mM caffeine in the presence of 20 mM KCl to monitor total sarcoplasmic reticulum (SR) load. Confocal imaging was accomplished at a spatial/temporal resolution of ~ 650 μm /36 fps; xt line scan resolution was ~1300 μm/650 fps.
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