Measurement of Vibration Detection Threshold and Tactile Spatial Acuity in Human Subjects

Summary

Here, we present protocols to determine vibration detection thresholds and tactile acuity using psychophysical methods in man.

Abstract

Tests that allow the precise determination of psychophysical thresholds for vibration and grating orientation provide valuable information about mechanosensory function that are relevant for clinical diagnosis as well as for basic research. Here, we describe two psychophysical tests designed to determine the vibration detection threshold (automated system) and tactile spatial acuity (handheld device). Both procedures implement a two-interval forced-choice and a transformed-rule up and down experimental paradigm. These tests have been used to obtain mechanosensory profiles for individuals from distinct human cohorts such as twins or people with sensorineural deafness.

Introduction

Specialized mechanosensory receptors in the skin mediate the perception of vibration and grating orientation. Each mechanosensory receptor type is tuned to detect distinct features of tactile stimuli^1,2. This property provides the psychophysical basis for a differentiated assessment of mechanosensory function by using tests that deliver simple sinusoidal oscillations (vibration) or fine gratings. It is known that psychophysical thresholds for vibration perception are lower for high than low frequency vibration^3,4. Two types of rapidly adapting mechanoreceptors associated with the Meissner and Pacinian corpuscles primarily detect low (10-40 Hz) and high (100-200 Hz) frequency vibration stimuli, respectively⁵. It is thought that the psychophysical thresholds for vibration perception rely to a considerable extent on the activation of these two sets of mechanoreceptors at their best frequencies^6,7. Tactile spatial acuity is tested by the grating orientation task to determine the finest grating whose orientation can be discriminated by a subject^7-9. Merkel cell-neurite complex afferents are essential for detecting grating orientation^8,10. Interestingly, there has been considerable recent progress in our understanding of the molecular basis of how mechanoreceptors detect extremely small tactile stimuli¹¹. Mechanosensitive ion channels like Piezo2 and modulators like STOML3 have been directly implicated in the detection of fine tactile stimuli in touch receptors^12-15. There are already human patients identified with function altering mutations in the Piezo2 gene and it will be important to test whether such patients have touch deficits¹⁶.

The determination of the vibrotactile threshold relies on the delivery of a vibrating stimulus to the skin. Vibration detection thresholds vary with the vibration frequency. Vibration frequency to detection threshold curves were described by von Bekesey, Verillo, and Bolanowski, among many others, from the 1930s up to the 1990s^5,17. The devices in the early days were based on shakers and power amplifiers and several devices have been commercialized and there are a lot of variation as to choice of stimulator (testing frequency), the diameter of the probe in contact with skin, the use of a surround to limit the stimulation to one area of the skin, and the testing protocol that determines the threshold (see the following references for more detailed insight on the device and testing features)^18-20. Most devices usually test one site; however, there are new devices that use vertical displacement stimulators equipped with 2 probes to deliver vibratory stimuli with separation ranges that can be varied²¹. Also detection thresholds are measured based on vibration frequency or intensity discrimination with continuous stimulation; or with intermittent vibration stimuli with or without a masking stimulus. Therefore, we recommend that the reader be aware of the plethora of developments in this field.

Here, we describe the components of the device and a step-by-step guide on how to conduct psychophysical tests to estimate vibration detection threshold in human subjects. We then show how to assess tactile acuity manually by using the tactile acuity cube. We use a two-interval, forced-choice paradigm: the stimulus is always presented during one of two intervals and the subject has to indicate which interval has the vibration stimulus. We employ a transformed-rule up and down as the adaptive method that executes a threshold search by changing the stimulus intensity based on the performance during the test. Both psychophysical protocols can be used by investigators as a screening tool for evaluation of alterations in touch sensitivity.

Protocol

The testing protocol was approved by the Charité-Universitätsmedizin Ethics Committee.

1. Vibration Detection Threshold (VDT)

1. Device and Testing Protocol Assembly – Pretesting
   1. Assemble the components of the device according to Figure 1A. Place a smooth-surfaced board (40 cm x 80 cm) on a table. Place the brass bar on the board.
   2. Connect the piezoelectric actuator (vibration stimulator) to the controller unit.
   3. Connect the response box and the monitor device to the data acquisition system (see supplemental code file).
   4. Connect the data acquisition system to a computer (or laptop), and to piezo actuator controller unit.
   5. Screw the custom-made stimulating probe to the moving part of the piezoelectric actuator (for specifics of the probe see Materials).
   6. Mount the piezoelectric actuator with the probe on the balanced brass bar.
2. Testing Protocol
   1. Script a test protocol that implements the two interval forced-choice and transformed-rule up and down method. See supplemental code file for an outline of the script.
   2. Construct the waveform of the vibration stimulus as a sinusoidal wave and specify stimulus duration, rise and fall characteristics.
      1. Open software (e.g., LabChart). Select Setup > Stimulator.
      2. Choose a custom waveform and configure the stimulator options. Create 2 stimulus waveforms pertaining to each interval (stim1 and stim2).
         NOTE: The waveform stim1 is composed of 3 parts: a delay of 4 sec, followed by a sinusoidal wave of 1.8 sec, and a delay of 1.8 sec (no stimulus). The waveform stim 2 is composed of 3 parts: a delay of 4 sec, followed a delay of 1.8 sec (no stimulus), and by a sinusoidal wave of 1.8 sec.
      3. For the sinusoidal waveform, create new variable parameters for frequency and amplitude. Modify the sinusoidal wave function by inputting the following functions to the rise and fall.
         Rise waveform: (1-e^-bt)∙Amplitude∙sine(frequency), b = 9.1
         Fall waveform: (e^-bt)∙Amplitude∙sine(frequency), b = 9.1
      4. In the data panel of the lab chart, create the set of 35 voltage outputs pertaining to the 35 amplitude intensities (or levels) of the vibration stimulus. See Table 1.
   3. Set the starting/default amplitude of the vibration stimulus for the tested vibration frequency in the macro script for the testing procedure (see supplemental code file section).
3. Preparation and Training of Subjects – Testing Session
   1. Inform test subjects about the testing procedure and have them sign a written consent form. To ensure anonymity and fulfill data protection requirements, assign each participant a number.
   2. Seat the subjects comfortably in a quiet room at temperatures between 20-30 °C. Instruct them about the test in a simple and clear manner so that the subject knows what to expect during the testing.
   3. Place the arm of the subject on the board. Pad the little finger with medical dough to minimize movement. Place the brass bar on the board to position the probe on the little finger of the tested hand just below the nail bed. Ensure proper contact between the probe and the skin and adjust the position of the probe to a horizontal position using the water-level. Avoid skin contact with the edges of the circular flat probe.
      NOTE: This ensures that the flat surface of the probe applies around 30 g (0.3 N) to the skin surface. Sharp edges can lead to decreased detection thresholds. The heavy mass of the brass bar prevents the transmission of distracting oscillations from the surroundings to the device and minimizes dissipation of the applied sine wave.
   4. Prior to testing, get the participants familiarized with the setup. Depending on the testing frequency, present both an easy (level 23) and a hard (level 7) to perceive vibration stimulus by varying the amplitude until the subject perceives a vibration to ensure that the experimental procedure is understood.
   5. If necessary, reset the starting (default) amplitude at the selected frequency so that the subject can easily detect the stimulus when the testing protocol starts (see 1.2.3).
   6. Minimize interactions between subject and examiner during the test.
   7. Start the test by running the script that uses the two-alternative forced-choice procedure with the up-down adaptive method²².
      NOTE: Actions in 1.3.7-1.3.12 are automated by the program script.
   8. In each trial, randomly administer a vibration stimulus during one of the two intervals that are visually indicated to the test subject as "1" and "2" on the screen of the monitor (Figure 1B). Have the subject indicate if the first or second sequential intervals contained the vibration stimulus by pressing one of two buttons "1" or "2" on the response box. Let the subject make a guess if he or she is not sure when the stimulus is presented.
      NOTE: The forced-choice technique requires that the subject responds even when the vibration is not perceived.
   9. In a trial series, which consists of 6 to a maximum of 9 single trials, repeat the same vibration stimulus at one amplitude level at least six times consecutively. If the responses are all correct, reduce the stimulus intensity level (down rule) for the subsequent trial series.
   10. Based on the decision rule of the adaptive method (logical operators in script), grant the subject more trials to the same stimulus intensity if the subject makes errors in a trial series. Reduce the stimulus intensity if the stimulus is correctly identified in at least 5 trials, and incorrectly in less than 2 trials.
   11. Increase the stimulus level if the subject makes the 2 incorrect responses in a trial series; or, more than one incorrect response and fewer than 5 correct responses.
   12. Document the change in the direction of stimulus intensity, as reversal point. Change the stimulus intensity according to reversal point number: prior to the third reversal point by 4 intensity levels; at the 3^rd reversal point by 2 intensity levels; else by 1 level (for more detail see supplemental code file and Figure 4B).
   13. End the testing when the subject completes a total of 8 reversals.
   14. Calculate the VDT by taking the median of the stimulus amplitude value of the last 6 reversals.

2. Tactile Spatial Acuity Test

1. Determine tactile acuity with a two-alternative forced choice grating orientation test using the Tactile Acuity Cube (TAC). The TAC is comprised of 6 sides each containing a grating (bar and groove) whose widths are 0.75 mm, 1.25 mm, 1.75 mm, 3.0 mm, 4.5 mm, and 6.0 mm.
2. Seat subjects in a quiet room at temperatures between 20-30 °C and instruct on the task.
3. During the experiment, blindfold the test subjects using shielded eyeglasses. Place the dominant hand on a table with the palmar surface facing up.
4. On each trial, apply the TAC to the finger pad at one of two grating orientations: vertically (parallel direction) or horizontally (transverse direction) aligned to the long axis of the finger. Randomly choose the order of the grating orientation for each trial.
5. Apply the gratings of the TAC for 2 sec to the finger pad of the index finger so that the cube exerts its whole weight on the finger (233 g). Avoid pressing the TAC on the finger pad.
6. Ask the subjects to determine the orientation of the alignment before the cube is removed from their finger.
7. Avoid movement of the participant's finger because it might provide a cue to the orientation. Discard the trial if the experimenter senses that the finger has moved.
   NOTE: Beware that small finger movements might not be detected by the experimenter in the procedure.
8. Employ a two-down and one-up adaptive method in the staircase algorithm.
9. Start with the largest grating, 6.0 mm.
10. Decrease the grating width after two correct identification of the orientation (correct response).
11. Test the next, smaller width and continue with the stepping rule until the subject makes an incorrect response and document the grating width as a reversal point.
12. Increase the grating width stepwise again until the two orientations of a width are determined correctly again.
13. End the test after completion of thirteen reversals.
14. Calculate the tactile grating orientation threshold by taking the median of the grating widths of the last 10 reversals.

Results

The piezoelectric actuator provides the vibration stimulus to the subject. The vibration stimulus has a total duration of 1.8 sec and is presented only once during a trial during the first or second interval (Figure 2A). The rise and fall time at the onset and offset of the stimulus is determined by the functions (1-e^-bt)∙Amplitude∙sine(frequency), and (e^-bt)∙Amplitude∙sine(frequency), respectively, where b is set at 9.1. The rise and fall time at on...

Discussion

The techniques used to evaluate VDT vary as to device specifications, hardware, and testing protocols. The International Organization for Standardization specifies the methods and procedures to analyze and interpret vibrotactile thresholds including recommendations for the various components of a vibrometer (ISO 13091-1 and 2^26,27) The described testing system abides to the relevant ISO recommendations for testing frequency range (4-125 Hz), method (variant of up-down staircase and forced choice), probe size (...

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Piezo actuator	Physik Instrument, Germany	P-602.1L	The linear piezoelectric actuator, with integrated position sensor and motion amplifier, contains a piezoceramic material that elongates and contracts when voltage is applied. The piezoelectric actuator travels up to100 µm. The actuator is equipped with a flexure guide that ensures straight motion without tilting or lateral offset. The displacement is linear and calibration is done and checked by the manufacturer. It is recommended that on-axis movement of the probe be checked under the microscope. According to the manufacturer, the stimulus amplitude dampens by less than 20% at oscillating frequencies of 1000 Hz. This can be checked by using a force or displacement measuring device (e.g. force transducer from Kleindiek).
Piezo Amplifier/Servo Controller	Physik Instrument, Germany	E-665	E-665 amplifier/controller drives and controls the displacement of a low-voltage piezoelectric actuator in a system with sensor position feedback (SGS sensors). The servo-controller provides the option for closed loop operation. When applying sinusoidal and oscillating stimuli the amplitude signal deviates from the set amplitude starting from 500 Hz and reaches a maximum decrease of 20% at 1000Hz.
LabChart Software	ADInstruments, USA	LabChart 7, MLU60/8	Can create, store and run macro of the psychophysical testing algorithm.
PowerLab	ADInstruments, USA	PowerLab 4/35 PL3504	Data Acquisition Hardware. Used with LabChart software.
Brass bar	Custom-made		Bar made of pure brass, weighs 15.5 kg. When the peizoelectric actuator is mounted on the brass bar it should exert a force of 30 g weight on skin surface.
Monitor	Custom-made		To mark the 1st and the 2nd interval. The monitor indicates to the subject the time intervals during which the stimulus may be presented.
Response box	Custom-made		The subject indicates the interval at which stimulus occurred.
Board	Custom-made		Upper surface should be smooth (Plastic), lower surface made of foam to prevent stray vibration ot be transmitted to the stimulating pobe.
Probe	Custom-made		A flat circular probe with smoothed edges (thermoplastic material) attached to a screw head. The screw should be of appropriate size to be tightened directly to the moving part of piezoelectric actuator. Size of the probe can be according to preference; in our case, diameter 8.21 mm and surface area 52.9 mm2.
Labchart Script			Can be sent on request. See supplementary code file.
Tactile Acuity Cube	MedCore		The cube is comprised of 6 sides each containing a grating (bar and groove) whose widths are 0.75, 1.25, 1.75, 3.0, 4.5, and 6.0 mm.