Pain Perception

The subjects were blinded to the procedures and could not see the sites undergoing stimulation from their supine position in the scanner. They were told that the acupuncture performed at different acupoints with different techniques would generate different needling sensations. Tactile (touch) stimulation was performed prior to acupuncture when the subjects were still naïve to acupuncture as a sensory comparison for the acupuncture stimulation. Thus, the comparison stimulation also took into account expectation and its placebo effects. Tactile stimulation and acupuncture were both performed at the same acupoint in 16 subjects each for LI4 and ST36, and 13 subjects for LV3. Three of the 42 subjects received tactile stimulation and acupuncture at all three acupoints; the remaining 39 subjects received acupuncture to all three acupoints, but only the paired tactile stimulation to the first of their acupoints. Analyses comparing tactile stimulation to acupuncture stimulation were performed on the paired sensory – acupuncture datasets. Data from all 3 acupoints for each subject was used in the Spearman's correlation of intensities of sensations in acupuncture.
Acupuncture and tactile stimulation control was delivered to LI4 on the hand, LV3 on the foot and ST36 on the lower leg on the right in randomized order by an acupuncturist with over 25 years of clinical experience (JL). The individual's sensitivity to needle manipulation was pretested, aiming to elicit deqi sensations without noxious pain. The stimulation paradigm is depicted in Figure 1. The needle was rotated approximately 180° in each direction with even motion at the rate of 60 times/min for 2 min during M1 and M2. The needle remained in place during the rest periods R1, R2 and R3. Each procedure lasted a total of ten minutes. In order to avoid excess discomfort, the subject was instructed to raise one finger if any sensation reached the intensity of 7–8 on a scale of 1–10 and 2 fingers in case of any sharp pain. When so signalled, the acupuncturist would adjust the force of stimulation so that the sharp pain would disappear within a few seconds. The acupuncture stimulation procedure was performed twice for each acupoint. Sterile, one-time use only stainless steel needles were used for LV3 (0.20 mm diameter) and ST36 (0.22 mm diameter) (KINGLI Medical Appliance Co. Wuxi, China). Silver needles (0.23 mm diameter) were used for LI4 (Matsuka, Tokyo, Japan). Superficial tactile stimulation was performed by gentle tapping with a size 5.88 von Frey monofilament, a standard method of sensory stimulation, prior to acupuncture with needling. The purpose of this design was to explore how acupuncture sensations might differ from the sensations elicited by the conventional sensory stimulus of touch. At the end of each tactile stimulation or acupuncture procedure, the subject was questioned by another researcher in the team if each of the deqi sensations (aching, pressure, soreness, heaviness, fullness, warmth, cooling, numbness, tingling, dull pain), sharp pain or any other sensations occurred during the stimulation, and to rate its intensity on the scale of 1–10 (1–3 mild, 4–6 moderate, 7–9 strong, 10 unbearable).

Pain perception in male and female was assessed at the age of 13, 15, 17, and 20 months, using the hot plate test as described (Dere et al., 2014 (link)). Briefly, mice were placed on a preheated (55°C) metal plate (Ugo Basile Srl, Comerio, Italy) and the latency (s) to retract by jumping or licking of the hind paws was recorded. Mice were exposed for a maximum of 40 s, used as cut-off time, in case they did not show an aversive response to the heated plate.