Proof Paper Depressing Efffect of Eletroacupuncture Quiroz Et Al

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Metadata of the article that will be visualized in OnlineFirst

ArticleTitle Depressing effect of electroacupuncture on the spinal non-painful sensory input of the rat

Article Sub-Title

Article CopyRight Springer-Verlag Berlin Heidelberg(This will be the copyright line in the final PDF)

Journal Name Experimental Brain Research

Corresponding Author Family Name Quiroz-Gonzlez

Particle

Given Name Salvador

Suffix

Division Departamento de Acupuntura y Rehabilitacin

Organization Universidad Estatal del Valle de Ecatepec

Address Av. Central s/n, Esq. Leona Vicario, Col. Valle de Anhuac, Secc. A,

Ecatepec, Estado de Mxico, CP 55210, Mexico

Email [email protected]

Author Family Name Segura-Alegra

Particle

Given Name Bertha

Suffix

Division Facultad de Estudios Superiores, FES Iztacala

Organization UNAM

Address Mexico, CP 54090, Mexico


Author Family Name Jimnez-EstradaParticle

Given Name Ismael

Suffix

Division Departamento de Fisiologa, Biofsica y Neurociencias

Organization Centro de Investigacin y, Estudios Avanzados del IPN

Address Av. Instituto Politcnico Nacional 2508, Col. San Pedro Zacatenco, AP.

14-740, Mexico, DF, CP 07000, Mexico


Schedule

Received 13 February 2014

Revised

Accepted 12 April 2014

Abstract The aim of this study was to explore the effect of electroacupuncture (EA) applied in the Zusanli (ST36) and

Sanyinjiao (SP6) points on the N1 component of the cord dorsum potential (CDP) evoked by electrical

stimulation of the sural nerve (SU) in the rat. The experiments were performed in 44 Wistar rats (250300 g)

anesthetized with ketamine (100 mg/kg) and xylazine (2 mg/kg). A bilateral laminectomy was performed to

expose the L3 to S2 segments of the spinal cord. The SU nerve was exposed and placed on pairs of hook

electrodes for electrical stimulation. The N1-CDPs were recorded with three silver-ball electrodes located on

the dorsal surface of the L5 to S1 segments. Ipsilateral high and low EA stimulation (100, 2 Hz, 6 mA, 30 min)

induced a considerable reduction in the amplitude (45 5.6, 41 6.2 %) of the N1-CDP recorded at the L6

segmental level. Recovery of the N1-CDP amplitude occurred approximately 13 s after EA. Sectioning of

the saphenous and superficial peroneal nerves reduced the depressing effect provoked by the EA stimulation


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(18.7 1.3, 27 3.8 %). Similarly, sectioning of the posterior and anterior tibial, deep peroneal and

gastrocnemius nerves partially reduced the effect provoked by EA (11 1.5, 9.8 1.1, 12.6 1.9 %).

Intravenous picrotoxin (1 mg/kg) also reduced the action of low and high EA (23 4.8, 27 5.2 %). It is

suggested that EA stimulation depresses non-painful sensory pathways through the activation of specific

inhibitory pathways that receive modulatory actions from other sensory and muscle afferent inputs in the rat

spinal cord.

Keywords (separated by '-') Electroacupuncture - Cord dorsum potentials - Sural nerve - Spinal cord

Footnote Information


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Journal : Large 221 Dispatch : 19-4-2014 Pages : 9

Article No : 3965 LE TYPESET

MS Code : EBR-14-0106 CP DISK

1 3

Exp Brain Res

DOI 10.1007/s00221-014-3965-2

RESEARCH ARTICLE

Depressing effect of electroacupuncture on the spinal non-painfulsensory input of the rat

Salvador Quiroz-Gonzlez Bertha Segura-Alegra

Ismael Jimnez-Estrada

Received: 13 February 2014 / Accepted: 12 April 2014

Springer-Verlag Berlin Heidelberg 2014

stimulation (18.7 1.3, 27 3.8 %). Similarly, sectioning

of the posterior and anterior tibial, deep peroneal and gas-trocnemius nerves partially reduced the effect provoked by

EA (11 1.5, 9.8 1.1, 12.6 1.9 %). Intravenous picro-

toxin (1 mg/kg) also reduced the action of low and high EA

(23 4.8, 27 5.2 %). It is suggested that EA stimulation

depresses non-painful sensory pathways through the activa-

tion of specific inhibitory pathways that receive modulatory

actions from other sensory and muscle afferent inputs in

the rat spinal cord.

Keywords Electroacupuncture Cord dorsum potentials

Sural nerve Spinal cord

Introduction

Acupuncture is a therapeutic modality that emerged from

Traditional Chinese Medicine. The World Health Organi-

zation recommends the use of acupuncture for the treat-

ment of a wide variety of diseases (Zhang et al. 2014; Yin

et al. 2010; Barnes et al. 2008). A novel contemporary

form of acupuncture, electrical stimulation of acupuncture

points (APs), also known as electroacupuncture (EA), has

been widely used in both clinical and experimental stud-

ies (Vickers et al. 2012; Zhao 2008). Several studies have

demonstrated that EA exerts an analgesic effect on neuro-

pathic pain in rat models (Lau et al. 2008; Kim et al. 2004;

Huang et al. 2004; Hwang et al. 2002) and relieves acute or

chronic inflammatory pain (Kim et al. 2006; Zhang et al.

2004). There has been particular interest in determining the

mechanisms involved in the antinociceptive effect of EA

(Zhao 2008; Leung 2012). Acupuncture analgesia is a clear

manifestation of modulatory processes occurring at differ-

ent levels of the central nervous system (Zhang et al. 2014;

Abstract The aim of this study was to explore the effect

of electroacupuncture (EA) applied in the Zusanli (ST36)and Sanyinjiao (SP6) points on the N1 component of the

cord dorsum potential (CDP) evoked by electrical stimu-

lation of the sural nerve (SU) in the rat. The experiments

were performed in 44 Wistar rats (250300 g) anesthe-

tized with ketamine (100 mg/kg) and xylazine (2 mg/kg).

A bilateral laminectomy was performed to expose the

L3 to S2 segments of the spinal cord. The SU nerve was

exposed and placed on pairs of hook electrodes for elec-

trical stimulation. The N1-CDPs were recorded with three

silver-ball electrodes located on the dorsal surface of the

L5 to S1 segments. Ipsilateral high and low EA stimu-

lation (100, 2 Hz, 6 mA, 30 min) induced a considerable

reduction in the amplitude (45 5.6, 41 6.2 %) of the

N1-CDP recorded at the L6 segmental level. Recovery of

the N1-CDP amplitude occurred approximately 13 s after

EA. Sectioning of the saphenous and superficial peroneal

nerves reduced the depressing effect provoked by the EA

S. Quiroz-Gonzlez (*)

Departamento de Acupuntura y Rehabilitacin, Universidad

Estatal del Valle de Ecatepec, Av. Central s/n, Esq. Leona Vicario,

Col. Valle de Anhuac, Secc. A, CP 55210 Ecatepec,

Estado de Mxico, Mexico

e-mail: [email protected]

B. Segura-Alegra

Facultad de Estudios Superiores, FES Iztacala, UNAM,

CP 54090 Mexico, Mexico


I. Jimnez-Estrada

Departamento de Fisiologa, Biofsica y Neurociencias, Centro

de Investigacin y, Estudios Avanzados del IPN, Av. Instituto

Politcnico Nacional 2508, Col. San Pedro Zacatenco,

AP. 14-740, CP 07000 Mexico, DF, Mexico


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Quiroz-Gonzlez et al. 2014; Zhao 2008). The first conver-

gence of impulses originating from pain sites and acupoints

occurs in the spinal dorsal horn, where the neuronal noci-

ceptive responses appear to be depressed by both pre- and

post-synaptic inhibition during EA stimulation (Zhao 2008;

Li et al. 1993). Although the effectiveness of EA for pain

control has been demonstrated experimentally in animal

models, little is known about the possible effects exerted byEA on the cutaneous non-painful sensory input in the spi-

nal cord. Electrical stimulation of cutaneous nerves induces

the activation of sensory neurons located in the dorsal

horn of the spinal cord (Bernhard 1953; Willis et al. 1973;

Coombs et al. 1956), which produces cord dorsum poten-

tials (CDPs) consisting of clearly defined components. The

first observed component in the CDPs is the afferent volley

(AV), which is caused by the electrical activation of low-

threshold afferent fibers (Coombs et al. 1956). This is fol-

lowed by one negative component (named N1-CDP), which

is generated by the monosynaptic activation of the dorsal

horn neurons via the Aafferents or non-painful fibers inthe cutaneous nerves (Bernhard 1953; Willis et al. 1973;

Coombs et al. 1956). Subsequently, a long-lasting positive

component occurs, named the P wave, which is ascribed to

the current flows generated during the presynaptic depolari-

zation of afferent fibers (PAD) and presynaptic inhibition

(Rudomin and Schmidt 1999). The present study aimed

to analyze the effect of EA stimulation on the non-painful

sensory neurons at the level of the spinal cord. To achieve

this goal, we recorded the N1-CDP evoked by cutaneous

SU nerve stimulation and during EA stimulation. In a sec-

ond series of experiments, we evaluated how this effect is

modified by sectioning cutaneous and muscular nerves.

Because it is amply recognize that GABAergic mechanism

is involved in the modulation of primary afferent depolari-

zation (PAD) of cutaneous and muscular nerves, we ana-

lyzed the possible effect of picrotoxin (PTX) on the actions

of the EA. Some of these observations have been published

previously in abstract form (Quiroz-Gonzalez et al. 2013).

Methods

Animals

Male Wistar rats (n = 44) weighing 250300 g (8

10 weeks old) that were obtained from the animal house

of our institution were used. All animals had free access to

water and were housed under identical environmental con-

ditions of light and dark cycles (12:12 h) and temperature

(2224 C). All experiments were carried out in accordance

with the guidelines of the Mexican Official Norm (NOM-

062-ZOO-1999) and in accordance with the National Insti-

tutes of Health Guide for the Care and Use of Laboratory

Animals (NIH Publication No. 8023, revised in 1978). The

study protocol was approved by the institutional bioethical

committee for Care and Handling of Laboratory Animals

(Protocol 0267-05, CINVESTAV).

Surgical procedures

Initially, the animals were anesthetized with an intraperito-neal injection of a mixed solution of ketamine (100 mg/kg)

and xylazine (2 mg/kg). This injection was supplemented

every hour by additional doses of ketamine (50 mg/kg),

applied in the same low abdominal region to ensure that an

adequate level of anesthesia, defined as the absence of with-

drawal reflexes. We used ketamine as an analgesic because

it has been demonstrated that it did not significantly reduce

the response of spinal dorsal horn neurons to low-threshold

afferent inputs in the intact animal, but in contrast suppress

noxiously evoked activity of wide dynamic range neurons

(Collins 1986). Subsequently, the femoral vein was exposed

and cannulated for the administration of a solution of pic-rotoxin (PTX, 1 mg/kg of body weight; Quirz-Gonzalez

et al. 2012). A bilateral laminectomy was performed in the

lumbosacral enlargement (from the L4 to S1 segments) of

the spinal cord. Several nerves of the right hind leg were

carefully exposed and prepared for stimulation and/or

sectioning: the main branch of the sural (SU), superficial

peroneus (SP), tibial (TA), saphenous (SA) deep peroneus

(DP) and gastrocnemius (GS) nerves and the rats were then

secured in a stereotaxic frame. The surgical procedure for

sural nerve dissection on the hind leg was carefully made

to avoid the possible damage of the Zusanli (ST36) and

Sanyinjiao (SP6) acupoints. The animals body temperature

was monitored using a thermal probe located in the back

muscles and connected to an automatic feedback control

unit and a heating blanket to maintain the animals body

temperature at 37 C. The skin flaps and muscle around the

exposed tissues were raised and tied to a metal stereotaxic

frame to form a pool, which was filled with warm mineral

oil to prevent the tissues from drying.

SU nerve stimulation

The central end of the sural (SU) nerve in the left hind

limb was mounted on a pair of stimulating silver hooks

connected to an isolated-current-pulse generator (Isoflex

D 4030), and the CDPs were produced in all experiments

by single square-current pulses (0.05 ms duration; at 1 Hz)

of graded intensity. The pulses were monitored by record-

ing the voltage drops across a resistor (1,000 ) that was

placed in the current return path. The electrical threshold

(1 T) was established as the minimum stimulus strength

(usually between 0.1 and 0.13 mA) necessary to cause a

discernible CDP response on the surface of the spinal cord.

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Electroacupuncture stimulation

Pairs of stainless-steel acupuncture needles (0.25 mm in

diameter and 5 mm in length) were inserted perpendicu-

larly at a depth of 3 mm at the Zusanli (ST36, 5 mm lateral

to the anterior tubercle of the tibia) and Sanyinjiao acu-

points (SP6, 3 mm proximal to the medial malleolus, at the

posterior border of the tibia). These acupoints are tradition-ally used in acupuncture-induced analgesia for the treat-

ment of several pain syndromes (Zhang et al. 2003; Huang

et al. 2002). To disclose the region-specific effect of EA,

we stimulate two non-acupoints (NAP) nearby to ST36 and

SP6, the first located 2 mm lateral to ST36 and the second

3 mm posterior to SP6. The transpositional method (Yin

et al. 2008) for rats was used to determine the acupoint

locations. The needles were connected to an isolated-cur-

rent- pulse generator (Isoflex D 4030), and square trains of

high or low frequency (100 or 2 Hz, respectively) and vari-

able strength current pulses (16 mA, 0.1 ms) were applied

in the acupoints located both contralateral and ipsilateral ofthe sural nerve stimulation for a total of 30 min.

CDP recording

Chlorinated silver-ball electrodes were placed on the dorsal

surface of several segments in the lumbosacral spinal cord

(L3S2) to record the CDPs (Fig. 1), and the corresponding

reference silver electrodes were inserted into the adjacent

paravertebral musculature. Each recording pair of elec-

trodes was connected to an individual low-noise, high-gain

differential amplifier (Grass, model P511; band-pass filters

were set at 0.310 kHz). The resulting recordings were dig-

itized, averaged (n=40 samples at 1 Hz), and stored in a

digital computer using a specially designed software (built

in the Lab-VIEW environment). The peak amplitude of the

N1-CDPs was measured and subsequently analyzed.

Data analysis

All statistical analyses were performed using the Graph-

Pad Prism (version 4) software. Data were expressed as the

mean standard deviation. A two-way analysis of variance

test for multiple comparisons followed by a Bonferroni cor-

rection was used to determine the differences in the ampli-

tudes of the N1-CDPs produced by SU nerve stimulation

and those produced by EA stimulation on ST36 and SP6.

Repeated measure ANOVA followed by Newman-Keuls

posthoc test was used to determine the differences between

the amplitude of control SU N1-CDPS and the amplitude

of the conditioned SU nerve responses evoked during EA

stimulation. The differences were considered significant at

p< 0.05.

Results

CDPs provoked by SU nerve and EA stimulation

Single electrical pulses applied to the SU nerve (34 T,

at 1 Hz) produced N1-CDPs of a relatively large amplitude

(Fig. 1a). The largest N1-CDP occurred at the L6-segmen-

tal level and gradually decreased in amplitude in the adja-

cent rostro-caudal spinal segments (L5, L4 and S1, respec-

tively, in Figs. 1a, 2a, 3a). Similar recordings were obtained

in 6 other experiments. This rostro-caudal distribution of

the CDPs agreed well with previous reports (Gonzlez et al.

2011). Single electrical current pulses applied to the ST36

and SP6 acupoints (34 T, at 1 Hz) produced N1-CDPs

with largest amplitude at the L5 and L6 segmental levels,

decreasing in amplitude in L4 and S1 spinal segments. The

N1-CDPs produced by SU nerve stimulation showed a sim-

ilar rostro-caudal distribution than those produced by EA

applied to ST36 and SP6, but they are smaller in amplitude

(p< 0.05; Fig. 1a, b).

Fig. 1 Experimental arrangement for the N1-CDP recording: aaver-

age N1-CDP produced by SU nerve stimulation and EA stimulation

recorded at L4 to S1 spinal cord segments. bGraphs illustrating the

longitudinal distribution on the spinal segments of the N1-CDP aver-

ages amplitudes (n=7) produced by SU nerve and EA stimulation at

ST36 and SP6. (*p< 0.05)

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Effect of high frequency EA stimulation on the amplitude

of the N1-CDP

The N1-CDP recorded on the surface of the L5 to S1 seg-

ments gradually decreased in amplitude when the high

frequency (100 Hz) EA stimulation strength was progres-

sively increased (from 1 to 6 mA; Fig. 2a, b). The maximal

depression of the N1-CDP (45 5.6 %) was observed at

the L6 segment (Figs. 2a, b, 3a, b) and occurred immedi-

ately upon and during EA stimulation (30 min). In most

Fig. 2 Inhibition of the

N1-CDP by high (100 Hz) EA

stimulation on ST36 and SP6:

aaverage N1-CDP produced

by SU nerve stimulation and

recorded in several spinal

segments (L5S1) before EA

(control) and during ipsilateral

or contralateral EA stimulation

at different stimulus intensities

(16 and 6 mA, respectively).

bGraphs illustrating the mean

reduction (n=11) in the

N1-CDP component recorded

in the S1L5spinal segments

during a 100 Hz ipsilateral EA

stimulation.Asterisksindicate

significant differences between

N1-CDP responses produced by

SU nerve stimulation before EA

and during high EA stimulation

(*p< 0.05 and **p< 0.01)

Fig. 3 Time course of inhibi-

tory effect of EA on N1-CDP

component: aCDP recording

showing that after EA, there

was a recovery of the N1-CDP

component produced by SU

nerve stimulation in several

spinal segments (S1L5).

bGraphs illustrating the

changes in the percent ampli-

tude of the N1-CDPs produced

before (control), during and

after EA. Asterisksindicate

significant differences between

N1-CDP responses produced

by SU nerve stimulation before

EA and during posthigh EA

stimulation (*p< 0.05 and

**p< 0.01)

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cases, the increment in intensity of EA stimulation (above

6 mA) did not increase the magnitude of the depression at

the different segments analyzed. Contralateral EA stimu-

lation (6 mA) and NAP stimulation had no effect on the

N1-CDP (Fig. 2a, b). After removal of the EA stimulus,

recovery of depressive actions on the N1-CDP occurred

within approximately 12 s (Fig. 3a, b).

Effect of low frequency EA stimulation on the amplitude

of the N1-CDP

30 min of low frequency EA stimulation (2 Hz) induce the

reduction in amplitude of the N1-CDP (41 6.2 %), but

only when the N1-CDP caused by EA stimulation occurred

at 100 ms before the N1-CDP produced by SU nerve stimu-

lation (Fig. 4B, C, D). No statistical differences were found

when the N1-CDP caused by EA stimulation occurred after

N1-CDP produced by SU nerve stimulation (Fig 4E, F).

Effect of nerve sectioning on the depressive action of EA

Sectioning the sensory saphenous and superficial pero-

neal nerves (Fig. 5a, b) induced a significant reduc-

tion in the depressing effect provoked by EA stimulation

on the SU-evoked N1-CDP (18.7 1.3 %, n= 7 and

27 3.8 %, n = 7, respectively). Meanwhile, section-

ing of the tibial, deep peroneal and gastrocnemius nerves

(Fig. 5a, b) reduced the depressing effect provoked by EA

on the N1-CDPs but to a lesser extent (11 1.5 %, n=7;

9.8 1.1 %, n=7; and 12.6 1.9 %, n=7, respectively).

The effect of PTX on the depressive action of EA

In order to analyze the possible GABAergic mechanism on

the depressive actions of EA on the SU-evoked N1-CDP,

systemic injection of a GABAA antagonist, picrotoxin

(PTX, 1 mg/kg) was delivered. As shown in Fig. 6D, F, the

application of PTX reduces the depressive actions of low

frequency EA (23 4.8 %, n=7), as compared with con-

trol recording (Fig. 6A, F). Similar reductions were found

(27 5.2 %, n=7) on the depressive actions of high fre-

quency EA stimulation (Fig. 6E, F). Intravenous saline

vehicle does not produced any effect on the EA conditioned

depression of the SU-evoked N1-CDP.

Discussion

It is well established that the N1-CDP generated by the

electrical stimulation of cutaneous nerves is produced by

the monosynaptic activation of dorsal horn neurons located

in the Rexeds laminae III to VI via A afferent fibers or

low-threshold cutaneous afferent fibers (Bernhard 1953;

Willis et al. 1973; Coombs et al. 1956). In our study, stimu-

lation of the sensory SU nerve and EA at the ST36 and SP6

acupoints provoked N1-CDPs that were simultaneously

Fig. 4 Inhibition of the SU N1-CDP by low (2 Hz) EA stimula-

tion applied on the ST36 and SP6 acupoints: a averaged N1-CDP(n=16 recordings) produced by SU nerve stimulation and recorded

in L6 spinal segment before EA stimulation, bduring EA (EA-CDP)

applied 70 ms previously to the stimulus of the SU nerve, c40 ms, d

20 ms, and e30 ms after SU-evoked N1-CDP (ASU-CDP). fGraphs

illustrate averaged (SD) percent reduction values of the N1-CDP

component recorded on the L6 spinal segment in nine animals, duringa 2 Hz ipsilateral EA stimulation. Asterisksindicate significant differ-

ences between N1-CDP responses produced by SU nerve stimulation

before EA and during low EA stimulation (*p< 0.05 and **p< 0.01)

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Fig. 5 Reduction in the

effect of EA at 100 Hz on the

N1-CDP component by section-

ing nerves: aCDPs recording

showing the effect of nerve sec-

tioning on the depressive action

evoked in N1-CDP by high EA

stimulation (100 Hz).

bGraphs illustrating the

changes in the percent ampli-

tude of the N1-CDPs produced

by EA and abolished by nerve

sectioning (n=7 animals per

experimental procedure). SP

superficial peroneous; SAsaphe-

nous; TAtibial; GSgastrocne-

mius soleus nerves. Asterisks

indicate significant differences

between control N1-CDP

responses evoked by SU nerve

stimulation and during EA

stimulation before and after the

sectioning of cutaneous and/or

muscular nerves (*p< 0.05 and

**p< 0.01)

Fig. 6 Effect of picrotoxin (PTX; 1 mg/kg) on the depressive effect

of EA on the N1-CDP: a averaged N1-CDP produced by SU nerve

stimulation and recorded in L6 spinal segment before EA stimula-

tion, bduring low frequency EA (2 Hz) applied 40 ms previously to

the stimulus of the SU nerve, c with high frequency EA (100 Hz),

d The effect of intravenous injection of PTX on the low frequency

EA conditioned depression of the SU-evoked N1-CDP, ePTX under

high frequency EA conditioned depression, f, glow or high frequency

EA+ vehicle intravenous administration, hgraphs illustrate averaged

(SD) percent reduction values of the N1-CDP component recorded

on the L6 spinal segment, during a 2 Hz EA stimulation (7 animals),

100 Hz EA stimulation (7 animals) intravenous PTX (7 animals),

and saline vehicle control (3 animals). Asterisks indicate signifi-

cant differences between N1-CDP responses produced by SU nerve

stimulation before EA, during EA stimulation and intravenous PTX

(*p< 0.05 and **p< 0.01)


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recorded on several segments of the lumbosacral enlarge-

ment (L4 to S1). The SU nerve evoked N1-CDP with the

largest was recorded on the L6 segment, and it is consist-

ent with those reported in other studies (Willis et al. 1973;

Gonzlez et al. 2011). Meanwhile, the largest N1-CDPs

evoked by EA were recorded at the L5L6 spinal segments.

These observations may suggest that the sensory inputs

activated by sural nerve and EA acupoint stimulation prob-ably share spinal pathways which probably interact synap-

tically at the spinal cord level. According to the later, we

also found that conditioning low and high frequency EA

stimulation depressed the N1-CDPs produced by SU nerve

stimulation. In contrast, EA stimulation on non-acupoint

sites does not evoke significant changes in the SU nerve

evoked N1-CDP, suggesting a specific acupoint effect of

EA. It is proposed that EA reduces the activation of dorsal

horn neurons provoked by low-threshold cutaneous afferent

fibers by the activation of specific sensory pathways in the

spinal dorsal horn of the rat.

Transmission of non-nociceptive and nociceptive infor-mation via primary afferents is modulated at the first spinal

relay by highly complex processes (Rudomin and Schmidt

1999; Le Bars 2002; Besson and Chaouch 1987). It has

been accepted that electrical stimulation of primary affer-

ent fibers effectively modulates the synaptic efficacy of

the same and/or other afferent fibers in the spinal cord (De

LaTorre et al. 2009; Rudomin and Hernandez 2008). Elec-

trophysiological evidences have shown that electrical stim-

ulation of Afibers may depress nociceptive activation of

spinal dorsal horn neurons for short periods of time (Chung

et al. 1984a, b). Moreover, both brief electrical stimulation

of afferent C-fibers and prolonged high frequency burst

stimulation of the sciatic nerve at Afiber strength produce

long-term depression (LTD) of C-fiber-evoked field poten-

tials (Liu et al. 1998). In addition, LTD of synaptic trans-

mission in substantia gelatinosa neurons can be induced

by low frequency stimulation of primary A-afferent fibers

(Sandkuhler et al. 1997).

Electroacupuncture stimulation also provokes consider-

able changes in the neuronal activity evoked by peripheral

nerve stimulation. Kim et al. (2011) showed that EA pro-

duced a significant reversal of enhanced evoked responses

of the deep dorsal horn (lamina IVVII) neurons as well as

after discharges developed in ankle-sprained rats. The EA-

induced inhibition lasted for at least 30 min after the ter-

mination of EA. In other study, it was found that 2 Hz EA

induce LTD in the C-fiber-evoked field potentials recorded

within the spinal dorsal horn of rats with neuropathic pain.

In contrast, 100 Hz EA-induced long-term potentiation

(LTP) but LTD in control rats (Xing et al. 2007).

In the present study, we found a significant reduction in

the amplitude of the N1-CDPs evoked by SU nerve stimu-

lation during ipsilateral EA stimulation (46 mA) in several

spinal segments, particularly at the L6 segment of the rat

spinal cord. The ST36 acupoint receives sensory innerva-

tion from saphenous, superficial peroneal, and lateral sural

cutaneous nerves and motor innervation from the deep per-

oneal and anterior tibialis nerves (Zhou et al. 2010), while

the SP6 acupoint receives sensory innervation from the

saphenous, sural and medial crural nerves and motor inner-

vation from the tibial nerve (Zhou et al. 2010). The spinalprojection of these nerves showed a considerable overlap,

particularly at the L5L6 segmental level, even though they

innervate different hind limb areas (Maslany et al. 1992;

Panneton et al. 2005). It thus seems reasonable to expect

that during EA stimulation, the sensory and motor inner-

vation of the acupoints are activated and that the highest

effect of EA is observed at the L6 segment.

It is known that the acupuncture effect may occur in a

gradual manner and last for a long period of time (Zhao

2008; Leung 2012). Several lines of evidence suggest that

neurotransmitters and endogenous opioids are involved in

the depressive effect of EA on spinal nociceptive neuronsand that they participate in the analgesic effect of acupunc-

ture (Zhao 2008; Leung 2012). In the present study, the

depressive effect of EA on the N1-CDP component showed

a fast onset (


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pathways (Rudomin and Schmidt 1999) or including to the

accumulation of potassium ions in the spinal cord (Kremer

and Lev-Tov 1998).

We also found that the depressive effects evoked by EA

were partially abolished by the sectioning of cutaneous and

muscle nerves that innervated the ST36 and SP6 acupoints.

The changes produced by sectioning of cutaneous nerves

were larger than those produced by the section of musclenerves. According to this evidence, it could be suggested

that specific heterosynaptic inhibitory pathways receiving

sensory and muscular inputs could be involved in the effect

of EA on low-threshold sensory pathways.

It is well known that the N1-CDP is produced by a

groups of sensory neurons that receive cutaneous large

A, afferent fibers (Bernhard 1953). These spinal neurons

located in the laminae III-VI are responsible to transmit

fine touch, vibration, propiocepcion to supraspinal cent-

ers (Willis et al. 1973). It may be suggest that EA at low

or high frequency affects the transmission of these differ-

ent sensory modalities at the spinal cord level and couldhave some implications in the process of the information

in the somatosensory cortex. Several lines of evidence

have hypothesized that large diameter sensory fibers play a

major role in the pathogenesis of some types of neuropathic

pain (Devor 2009; Campero et al. 1998). Devor (2009)

has showed that dorsal root ganglion A afferents, which

normally signals touch and vibration, change their electri-

cal and neurotransmitter characteristics when they are sec-

tioned (axotomized). Such condition seems to switch the

sensory input of A afferents from non-painful to painful

signals (phenotypic switching), triggering, and maintain-

ing central sensitization.

Since it has been shown that EA stimulation exerts anal-

gesic and antinociceptive effects by modulating the activ-

ity of spinal dorsal horn neurons and the experimental

evidence obtained in this study indicates that high and low

frequency EA stimulation also affect low-threshold non-

painful sensory pathways at the spinal cord level in the rat,

it could be proposed that the depression of low-threshold

cutaneous pathways is involved in the reduction in neuro-

pathic pain produced by EA stimulation. However, further

studies are necessary to analyze this possibility by analyz-

ing the effect of EA on low-threshold sensory pathways in

an animal model of neuropathic pain.

In conclusion, the present study showed that EA stim-

ulation depressed non-painful sensory spinal pathways

through the activation of specific inhibitory pathways that

receive modulatory actions from sensory and muscle affer-

ent inputs in the rat spinal cord.

Acknowledgments We thank American Journal Experts for edit-

ing the English of this text, Jos Carlos Guadarrama Olmos for tech-

nical assistance and to Enrique Velazquez and Porfirio Reyes for

their programming assistance. This work was partially supported

by fellowships granted to I. Jimnez-Estrada and B. Segura-Alegra

from the Sistema Nacional de Investigadores. S. Quiroz-Gonzalez

was partially supported by PROMEP (No. 103.5-13-6729) and

SNI-CONACYT.

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