The Impact of Tenderness on Prefrontal Cortical 1 Hemodynamics: An Objective Measurement of Wrist-ankle 2 Acupuncture Analgesic Bracelet

9 Background: Determining an objective measure for pain is one of the most significant 10 challenges in neuroscience and clinical medicine. The response of the cerebral cortex 11 to noxious stimuli/pain can be detected by functional near infrared spectroscopy 12 (fNIRS). The study aimed to examine the law of pain relief and cerebral blood volume 13 dynamics based on the wrist-ankle acupuncture (WAA) analgesic bracelet, and 14 determined the reliability of the cortical activation patterns as objective measurements 15 of pain. 16 Methods: Tenderness was performed on the left point Jianyu of volunteers (age, 23.9 17 ± 1.9 years, mean ± SD) suffering from the cervical-shoulder syndrome (CSS) as tasks. 18 A transcutaneous electrical nerve stimulation analgesic bracelet based on WAA in 19 efficacy of an analgesic bracelet based on WAA in TCM for the first time, and provides 40 a new research direction for further pain relief.


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Pain, according to the International Association for the Study of Pain, is a 'subjective' 47 Participants 105 Previous studies have suggested that sex-related hormones could be confounding in the 106 relationship between pain and the analgesic response [17,18]. Consequently, only men 107 were selected as participants in this study. The subjective analgesic effect of the 108 analgesic bracelet used in this study has been confirmed [19]. Given that the analgesic 109 bracelet is in the stage of applying for medical device registration, the experimental 110 data comes from the small-scale participants recruited via an announcement addressed 111 to faculty members and students affiliated with University of Shanghai for Science and 112 Technology. They are all social members of different ages, and the data is somewhat 113 representative. All participants are right-handed to avoid any variation in functional 114 response due to lateralization of cerebral function. Fifteen right-handed with CSS males 115 aged 23.9 ± 1.9 years (mean ± SD) gave their informed consent after receiving 116 explanations on the study's significant points. All procedures were approved by the 117 Ethics Committee of ChiRCT. 118

Experimental devices 119
The tenderness experiment of point Jianyu was applied via a system typically used for 120 measuring muscle condition quickly. In consideration of safety, a tenderness meter 121 (OE-220: Ito Corporation, Tokyo, Japan) for quantitative analysis of perceptual pain 122 was used to deliver pain stimuli associated with CSS. The tenderness site was selected as the point Jianyu, which is related to CSS, determined by the mechanism of 124 symptomatic acupoint searching in TCM, shown in Fig. 1a.

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The analgesic treatment was applied through a transcutaneous nerve electrical 132 stimulation analgesic bracelet based on WAA in TCM. It is a miniaturized portable 133 non-invasive electrical stimulation device that can be worn on the wrist or ankle with 134 Velcro to relieve and even treat the human body's pain. Gold finger electrodes that 135 output electrical pulses are embedded in Velcro and directly contact the human skin. 136 The treatment site was selected as the upper 5 area on the left hand, which is related to 137 point Jianyu, and is determined by the mechanism of WAA in TCM (Fig. 1b). The 138 upper 5 area is located in the middle of the palm surface between the two most 139 prominent palmar longus tendons and the flexor carpi radialis tendons [20]. Due to the specificity of pain perception, the treatment parameters were adjusted themselves: 0-141 100 V, 0-100 Hz. 142

Experimental Procedure 143
The study was conducted in a laboratory shielded from interference. The fNIRS's 144 measurement sometimes results in hemoglobin changes due to factors unrelated to 145 cerebral activity, such as poor optical fiber contact caused by body movement or 146 changes in blood flow associated with muscle activity. Therefore, participants should 147 sit and stay as still as possible during the experiment. All the room lights were switched 148 off (light intensity controlled below 100 lx) to minimize signal contamination from the 149 ambient light sources. Participants were informed of the experimental procedures by 150 the researchers before the experiment. The experimental procedures were performed by 151 experienced TCM experts. 152 The experimental procedure was based on five sets, including three sets of tenderness 153 experiments and two sets of treatments, with a 1-minute recovery time between sets, 154 shown in Fig. 2. A set of tenderness experiments was composed of three blocks of 20 155 seconds of tenderness, followed by 20 seconds of rest (no stress). A set of treatments 156 refers to a 5-minute electrical stimulation treatment with an analgesic bracelet. During 157 each experiment, the participants were asked to sit down on a comfortable chair, relax, 158 close their eyes, and avoid major body movement. The TCM experts stood on the left 159 side of the participant. A preparatory period of 1 minute was set prior to data collection. 160 The procedure lasted 22 minutes in total.
In the tenderness experiment, the participants' left point Jianyu was pressed vertically 162 until the instrument emits a beeping sound with the tenderness meter, each time for 20 163 seconds. Normally, a sense of pressing pain is felt upon the application of a test point. 164 Therefore, the Visual Analogue Scale (VAS; range: 0 -100 mm) was used to evaluate 165 the intensity of the pain, and results showed a VAS level of 27 ± 6 mm (mean ± SD), 166 which is indicative of a mild pain stimulus [21,22]. 167 In the analgesic treatment, the two output golden fingers of the analgesic bracelet were 168 placed on the upper 2 area of the participants' left hands for 5 minutes. The bracelet 169 outputs 2 Hz and 100 Hz pulses in sequence every 1 minute. The participants adjusted 170 their parameter settings until they could bear the maximum intensity without pain 171 through the intelligent terminal. The participants' analgesia parameters were set to 40 172 ± 2.1 V (mean ± SD). 173 were used for the analysis, due to its higher signal-to-noise ratio [24]. The measurement 185 area was the prefrontal cortex region, as it was expected to obtain a hemodynamic 186 response in this cortical region. Therefore, artifacts related to blood influx to the 187 temporal muscle most likely did not affect our data. The probe targeting the prefrontal 188 cortex was placed so that the array at its bottom coincided with the line of T3-Fpz-T4 189 (International 10-20 system reference points) [25]. Probes were placed on the right side 190 of the forehead of each participant, and the end of the probes was adjusted to align with 191 the middle of the forehead-to cover position Fp1, Fp2 of the international 10-20 EEG 192 system (in Fig. 3). The configuration for this experiment was nine probes of 12 channels. 193

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The vertical axis shows the measured values (unit: mM mm), and the horizontal axis shows time (unit: 243 second). In this study, the analysis was conducted on Oxy-Hb levels obtained from 12 channels placed 244 over the right hemisphere's prefrontal cortex. The numbers showed represent channel numbers.

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Additionally, fNIRS cannot measure the exact optical path length, and the measured 246 value obtained can only reflect a relative change from the baseline condition rather than 247 the absolute concentration. fNIRS measurements cannot be compared between 248 participants in different states or regions within a participant. Thus, in the present study, 249 the Oxy-Hb concentration changes in each channel in three tenderness experiments by 250 calculating the differences between the rest and the task data were compared. Table 1  251 shows the mean ± SD of changes in Oxy-Hb levels during tenderness load, calculated 252 by the 12 channels of the right hemisphere. The mean ± SD of Oxy-Hb levels changes 253 in three tenderness were 0.50±1.22 mM mm, 0.28±1.36 mM mm, 0.09±0.81 mM mm, 254 respectively. The Oxy-Hb concentration changes in three tenderness experiments 255 gradually decreased, and the changes in channels 2, 5, and 11 even reduced to a negative 256 value during the third tenderness. CH6 showed the maximum difference in blood Oxy-257 Hb concentration in three tenderness experiments. Further, paired t-tests were 258 performed on the changes in Oxy-Hb levels in the right hemisphere's 12 channels. After 259 the initial treatment and intensive treatment with the analgesic bracelet, each targeted 260 channel's Oxy-Hb changes caused by three tenderness loads showed a decreasing trend, 261 and there were significant differences between the two pairs. 262  fMRI. However, fMRI equipment is bulky, and the test space appears to be closed, 294 which often causes tremendous psychological pressure on patients and affects the 295 evaluation results. Therefore, the multi-channel fNIRS, which is small in size and does 296 not affect the patients' daily activities, was used to evaluate pain and analgesia 297 objectively. Specifically, the effect of pain on the activation of the cerebral cortex is 298 determined by measuring the change in Oxy-Hb levels of the Jianyu point of CSS 299 patients during tenderness, which further provides a basis for cerebral blood volume 300 dynamics and cerebral cortex activation patterns as indicators for objective assessment 301 of pain. In addition, the efficacy of this therapy is first evaluated objectively by the 302 cerebral blood volume dynamics and the cerebral cortex activation as pain assessment 303 indexes of CSS patients. 304 The ability of fNIRS to detect cerebral activation and hemodynamic response following 305 a stimulus involving tenderness of the point Jianyu was verified, as shown in Fig. 4 and 306 Table 1. As oxygen is delivered to neurons by hemoglobin, many neuroimaging studies 307 use increased cerebral hemodynamic response signal intensity as a cerebral activation 308 marker [35]. In the present study, the Oxy-Hb changes were used as a measure for 309 cerebral activation. Fig. 4 reveals that neural activity in the hemisphere (the perception, conduction, and processing of pain) caused by tenderness leads to increases in regional 311 cerebral blood flow and regional cerebral Oxygen metabolic rate. 312 Furthermore, since the 12 channels of the right hemisphere correspond to the following 313 anatomical cerebral regions: the frontopolar area (CH1, CH3, CH4, CH5), as well as 314 the dorsolateral prefrontal cortex (CH2, CH6, CH7, CH8, CH11), and the includes 315 frontal eye fields (CH9, CH10, CH12), paired t-tests (two-tailed) and Bonferroni 316 correction revealed a trend at these channels for Oxy-Hb to increase. The activation 317 maps shown in Fig. 5 illustrate the spatial distribution of activation among the 12 318 channels. The plots were conducted to demonstrate the activation area's consistency in 319 each set for the tenderness experiment. For the tenderness task, the activated cerebral 320 region is consistent within most individual participants' trials. Small inconsistencies 321 were found in some trials, but these can be attributed to the set-to-set variation present 322 in the fNIRS signals. In addition, the net increase in blood flow caused by the painful tenderness of the point 345 Jianyu showed a decreasing trend after the analgesic bracelet treatment, and the trend 346 was more evident after the intensive treatment, and even the net increase of CH2, CH10, 347 and CH11 decreased to a negative value. After applying an analgesic bracelet based on 348 the theory of WAA in TCM, the hemisphere's stress system secretes a large amount of 349 corticotropin-releasing factor, dynorphin and glutamic acid, and flows into the 350 bloodstream under the same tenderness. It was thereby weakening the hemodynamic 351 blood response. After intensive treatment, the release of dopamine in the limbic 352 dopamine system in the hemisphere is reduced. The body is in a low dopaminergic state, 353 which is clinically manifested as anhedonia and motivation or an intrinsically 354 significant decrease in natural enhancers, and the hemodynamic response is still 355 weakened. Therefore, this study not only supports the application of cerebral blood 356 volume dynamics and cerebral cortex activation patterns in the objective assessment of 357 pain, but also realizes the objective quantification of the evaluation of the analgesic 358 bracelet based on WAA. 359 To the extent that the analgesic bracelet influences clinical pain, the possibility for novel 360 treatment applications exists. For example, the analgesic bracelet appears to be effective 361 in treating cyclical frozen shoulder in men. It seems plausible that analgesic bracelet 362 therapy based on WAA in TCM would effectively treat other chronic pain conditions. 363 Further, if such treatments potentiate other analgesics' effects, this would provide a 364 more optimal therapeutic profile. The efficacy of analgesic bracelet therapy based on 365 WAA in TCM for the amelioration of chronic pain remains an important yet unexplored 366 area of research. 367 The participants in this study are all male. To some extent, this study reflects the 368 hemodynamic response of the cerebral cortex of young men to shoulder tenderness. The 369 large difference in the pain perception between males and females suggests that the 370 research on the hemodynamic response of different genders to the application of wrist-371 ankle acupuncture analgesia in the future.
Several limitations of this study should be noted, however. The study was conducted 373 on shoulder tenderness in patients with CSS aged approximately 20 years. However, 374 the diagnosis population of CSS is very wide, and patients of different ages may react 375 differently to tenderness. The participants in this study are all young people. Further, a 376 study examining the human hemisphere's proportions dedicated to processing sensory 377 functions reported that the primary somatosensory cortex area related to the body trunk 378 was smaller than that of the hand or face [38]. A stimulation involving pressure only 379 causes a slight change in cerebral blood flow. Thus, further studies are required to use 380 a more vital stimulation to point Jianyu, such as acupuncture, which may cause more 381 incredible cerebral blood flow changes. 382