Articles in PresS. J Neurophysiol (September 2, 2015). doi:10.1152/jn.00444.2015
1 2 3 4 5 6 7 8 9 10 11 12 13 14
Characterizing pinprick evoked brain potentials before and after experimentally-induced secondary hyperalgesia
Emanuel N. van den Broeke 1*, André Mouraux 1, Antonia H. Groneberg 2, Doreen B. Pfau 2, Rolf-Detlef Treede 2 and Thomas Klein 2 Affiliations: 1 2
Institute of Neuroscience, Université catholique de Louvain, B-1200, Brussels, Belgium Division of Neurophysiology, Center for Biomedicine and Medical Technology Mannheim (CBTM), Heidelberg University, 68167, Mannheim, Germany.
15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42
*Corresponding author: Emanuel N van den Broeke Institute of Neuroscience (IoNS) Faculty of Medicine Université catholique de Louvain B-1200, Brussels Belgium E-mail:
[email protected]
Keywords: capsaicin, secondary hyperalgesia, central sensitization, pinprick, evoked potentials.
1 Copyright © 2015 by the American Physiological Society.
43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64
ABSTRACT Secondary hyperalgesia is believed to be a key feature of “central sensitization” and is characterized by enhanced pain to mechanical nociceptive stimuli. Using EEG, the aim of the present study was to characterize the effects of pinprick stimulation intensity on the magnitude of pinprick elicited brain potentials (event-related potentials; ERPs), before and after secondary hyperalgesia induced by intradermal capsaicin in humans. Pinprick elicited ERPs and pinprick-evoked pain ratings were recorded in 19 healthy volunteers, using mechanical pinprick stimuli of varying intensities (0.25 mm probe applied with a force extending between 16 and 512 mN). The recordings were performed before (T0) and 30 minutes after (T1) intradermal capsaicin injection. The contralateral non-injected arm served as control. ERPs elicited by stimulation of untreated skin were characterized by (1) an earlylatency negative-positive complex peaking between 120 and 250 ms after stimulus onset (N120-P240) and maximal at the vertex, and (2) a long-lasting positive wave peaking 400-600 ms after stimulus onset and maximal more posterior (P500) which was correlated to perceived pinprick pain. After capsaicin injection, pinprick stimuli were perceived as more intense in the area of secondary hyperalgesia and this effect was stronger for lower compared to higher stimulus intensities. In addition, there was an enhancement of the P500 elicited by stimuli of intermediate intensity, which was significant for 64 mN. The other components of the ERPs were unaffected by capsaicin. Our results suggest that the increase in P500 magnitude after capsaicin is mediated by facilitated mechanical nociceptive pathways.
65 66 67
INTRODUCTION
68
Cutaneous tissue injury is often associated with the development of increased pain
69
sensitivity in the area of actual tissue injury (referred to as primary hyperalgesia) and in the
70
surrounding uninjured skin (referred to as secondary hyperalgesia).
71
A hallmark of secondary hyperalgesia is enhanced pain to mechanical nociceptive stimuli
72
(e.g. pinprick stimuli; Raja et al. 1984; Ali et al. 1996; Magerl et al. 1998). It can be induced
73
experimentally by activating nociceptors in a sustained and intense fashion, for example,
74
using intradermal injection or topical application of capsaicin, a substance that selectively
75
activates primary nociceptive afferents via the TRPV1 receptor (LaMotte et al. 1991; Magerl 2
76
et al. 1998; Magerl et al. 2001; Ziegler et al. 1999). Some studies suggest that secondary
77
pinprick hyperalgesia is primarily mediated by capsaicin-insensitive A-fibers, which include
78
high-threshold mechanoreceptors (HTM) and Type I A-fiber mechano-heat nociceptors
79
(AMH-I) (Magerl et al. 2001; Ziegler et al. 1999), and results from “central sensitization”
80
(Baumann et al. 1991; LaMotte et al. 1991; Latremoliere and Woolf 2009; Simone et al.
81
1991), which is defined by the International Association for the Study of Pain (IASP) as
82
“increased responsiveness of nociceptive neurons in the central nervous system to their
83
normal or subthreshold afferent input” (Loeser and Treede 2008).
84
Experimentally induced secondary pinprick hyperalgesia seems very similar to the observed
85
hyperalgesia in patients with pain following lesions of the peripheral and central nervous
86
system (Jensen and Finnerup 2014). Until now the measurement of pinprick hyperalgesia
87
relies predominantly on the subjective report of pain intensity. It would thus be of great
88
value to have a non-invasive clinical and research laboratory tool that is able to reliably
89
assess sensitization of mechanical nociceptive pathways. One of these tools could be the
90
recording of pinprick event-related brain potentials (ERPs).
91
Recently, Iannetti et al. (2013) recorded for the first time ERPs elicited by mechanical
92
pinprick stimulation before and after intradermal injection of capsaicin in the area of
93
secondary hyperalgesia, in healthy volunteers. The mechanical pinprick stimulation was
94
performed by quickly applying and removing a sharp-tipped probe (0.25 mm diameter). The
95
probe could slide freely inside a tube, handheld by the experimenter. A calibrated weight on
96
top of the load was used such that the normal force was 128 mN when the needle is
97
maintained against the skin. The stimulus elicited a biphasic ERP waveform (N120 and P240
98
waves) with latencies compatible with the conduction of myelinated Aβ or Aδ fiber 3
99
afferents. After capsaicin injection, they observed an enhancement of both the pain ratings
100
and the magnitude of the pinprick-evoked N120 wave.
101
However, only one force was used in this seminal study, leaving unexplored the possibility
102
that pinprick evoked potentials are modulated differently depending on the applied force,
103
both in untreated skin and in the area of secondary hyperalgesia. Therefore, the aim of the
104
present study was to characterize the ERPs elicited by different intensities of pinprick
105
stimulation, before and after inducing secondary hyperalgesia. Different weights were used
106
to deliver a range of stimulation intensities, extending between 16 and 512 mN. Secondary
107
hyperalgesia was induced by intradermal injection of capsaicin.
108
109
2. METHODS
110
2.1 Participants
111
Nineteen healthy volunteers took part in the experiment (9 men and 10 women; aged 19 –
112
29 years; mean ± SD age: 23.6 ± 2.3 years, 18 right handed, 1 left handed). Approval for the
113
experiment was obtained from the local Ethical Committee. All participants signed an
114
informed consent form.
115
116
117
118
4
119
2.2 Experimental design
120
Participants were comfortably seated in a reclining chair with their arms resting on armrests.
121
Two concentric circles were drawn on the left and right ventral forearm, with a diameter of
122
1.5 cm and 2.5 cm, respectively.
123
124
2.3 Intradermal injection of capsaicin
125
A 10 mM solution of capsaicin (40 µg capsaicin dissolved in 12.5 µl of 16% Tween80 in
126
normal saline; Magerl et al. 2001) was injected in the dermis, at the center of the circles
127
drawn on the ventral forearm of the non-dominant arm (Fig. 1). Immediately after the
128
injection, participants were asked to rate the magnitude of the capsaicin-induced pain every
129
10 sec. for the first minute and then every 30 sec. until 15 minutes post injection, using a
130
numeric rating scale (NRS) ranging from 0 (no pain) to 100 (most intense pain imaginable). A
131
placebo saline injection was not included in this study, as the aim was not to ascertain
132
whether capsaicin induces hyperalgesia, but rather to examine the relationship between this
133
well-known phenomenon and pinprick ERPs.
134
135
2.4 Pinprick stimulation
136
Pinprick elicited ERPs were recorded before (T0) and 30 minutes after (T1) capsaicin
137
injection, using six different intensities of mechanical pinprick stimulation: 16, 32, 64, 128,
138
256 and 512 mN. The pinprick stimuli were delivered to the capsaicin-treated and control
139
arm, within the 1-cm wide area defined by the two circles drawn on the forearms at a 5
140
distance ranging between 1.5 and 2.5 cm from the site of capsaicin injection on the treated
141
arm (Fig. 1).
142
For each intensity of pinprick stimulation and for each arm, a total of twenty stimuli were
143
administered in separate blocks. Stimulation of the control and capsaicin-treated arm was
144
performed in an alternating fashion (block-by-block design), but the capsaicin-treated arm
145
was always stimulated first. The order of the different stimulation intensities was balanced
146
across participants. The stimuli were delivered using a random inter-stimulus interval
147
ranging from 7 to 10 sec. To avoid sensitization of the stimulated skin, the target of the
148
pinprick stimulus was displaced by the experimenter after each stimulus.
149
The pinprick stimuli were generated by a set of six probes consisting of a cylindrical stainless-
150
steel 0.25 mm diameter flat-tip (uniform tip geometry) mounted on a plastic rod (MRC
151
Systems GmbH, Heidelberg, Germany). The plastic rod moved freely inside a polished hand-
152
held stainless steel tube. Each pinprick stimulus was delivered by applying the needle in
153
vertical direction onto the skin and moving the tube downwards and upwards with a total
154
duration of approximately one second. The normal force applied by the needle against the
155
skin (16, 32, 64, 128, 256 and 512 mN) was adjusted by placing inside the tube and on top of
156
the plastic rod, one of a set of six calibrated cylindrical weights.
157
Instead of applying the pinprick stimulus as fast as possible such as in the study of Iannetti et
158
al. (2013), the pinprick stimulus was applied and removed slowly, and maintained against
159
the skin for at least one second. The reason for doing so is that the actual force that is
160
applied onto the skin is dependent on the velocity of which the stimulus is delivered. To be
161
sure that the actual delivered force is the same as the planned one, the pinprick stimulus 6
162
should
be
applied
onto
the
skin
163
systems.de/englisch/products/pinprick.html).
in
a
slowly
fashion
(www.mrc-
164
165
2.5 Intensity of perception
166
The effect of capsaicin on the intensity of perception elicited by pinprick stimulation was
167
assessed by asking participants to rate the intensity of “pinprick pain” on a NRS ranging from
168
0 (no pain) to 100 (most intense pain imaginable). Participants were instructed to distinguish
169
the perception of pain (i.e. sharp or slightly pricking or burning sensation) from touch or
170
pressure by rating the stimulus > 0 (Rolke et al. 2006). Subjects were free to use integers as
171
well as fractions. Ratings of the intensity of perception were obtained 3 s after each
172
individual pinprick stimulus.
173
174
2.6 Electro-encephalography (EEG)
175
The EEG was recorded using five Ag-AgCl electrodes (Easycap, Brain Products GmbH,
176
Germany) mounted in an elastic electrode-cap and located at Fz, Cz, Pz, T3 and T4 according
177
to the International 10/10 system. Linked earlobes (A1A2) served as reference. Eye blinks
178
were recorded using a pair of surface electrodes placed at the upper and lower sides of the
179
right eye. Impedance was kept below 5kΩ for all leads. The signals were amplified and
180
digitized at a 200 Hz sampling rate using a BrainAmp system (Brain Products GmbH,
181
Germany). During the recording, participants were instructed to keep their gaze fixed on a
7
182
reference point at a distance of approximately 1 m at eye level, and to sit as still as possible
183
without making any movements.
184
The experiment was carried out with a modified set of pinprick probes (MRC Systems GmbH,
185
Heidelberg, Germany). The stimulators were equipped with an electrically conductive
186
connection from the needle tips via the internal sliding body to an external cable. As soon as
187
the probe came in contact with the skin, the electrical circuit detected the change in circuit
188
impedance and generated a trigger signal.
189
190
2.7 Data analysis
191
2.7.1 Intensity of perception
192
Intensity ratings of capsaicin- and pinprick-evoked pain were transformed into decadic
193
logarithmic values to obtain a secondary normal distribution. A small constant of 0.1 was
194
added to pain ratings to avoid a loss of zero values (Magerl et al. 1998).
195
To characterize the effect of capsaicin on the intensity of the percept elicited by pinprick
196
stimulation, a General Linear Model (GLM) repeated measures ANOVA analysis (Statistica
197
4.5) was performed on log-transformed pain ratings using three within-subject factors: time
198
(two levels: T0, T1), treatment (two levels: control arm, capsaicin arm) and stimulation
199
intensity (six levels: 16, 32, 64, 128, 256, 512 mN). The level of significance was set at p