Treede Timo Kirschstein, Wolfgang Greffrath, Dietrich Büsselberg ...

Timo Kirschstein, Wolfgang Greffrath, Dietrich Büsselberg and Rolf-Detlef 

Treede 

J Neurophysiol 82:2853-2860, 1999. 

You might find this additional information useful... 

This article cites 43 articles, 14 of which you can access free at: 

http://jn.physiology.org/cgi/content/full/82/6/2853#BIBL 

This article has been cited by 13 other HighWire hosted articles, the first 5 are: 

A multi-scale view of skin thermal pain: from nociception to pain sensation 

Y. J. Zhu and T. J. Lu 

Phil Trans R Soc A, 

February 13, 2010; 368 (1912): 521-559. 

[Abstract] [Full Text] [PDF] 

Heat-Induced Action Potential Discharges in Nociceptive Primary Sensory Neurons of Rats 

W. Greffrath, S. T. Schwarz, D. Busselberg and R.-D. Treede 

J Neurophysiol, 

July 1, 2009; 102 (1): 424-436. 


Heat Sensitization in Skin and Muscle Nociceptors Expressing Distinct Combinations of 

TRPV1 and TRPV2 Protein 

K. K. Rau, N. Jiang, R. D. Johnson and B. Y. Cooper 

J Neurophysiol, 

April 1, 2007; 97 (4): 2651-2662. 


TRPV1b, a Functional Human Vanilloid Receptor Splice Variant 

G. Lu, D. Henderson, L. Liu, P. H. Reinhart and S. A. Simon 

Mol. Pharmacol. , April 1, 2005; 67 (4): 1119-1127. 


Differential Effects of CB1 and Opioid Agonists on Two Populations of Adult Rat Dorsal 

Root Ganglion Neurons 

I. A. Khasabova, C. Harding-Rose, D. A. Simone and V. S. Seybold 

J. Neurosci. , February 18, 2004; 24 (7): 1744-1753. 


Medline items on this article's topics can be found at http://highwire.stanford.edu/lists/artbytopic.dtl 

on the following topics: 

Biochemistry .. Membrane Conductance 

Physiology .. Cation Channel 

Biochemistry .. Capsaicin 

Physiology .. Nociception 

Physiology .. Action Potential 

Chemistry .. Cations 

Updated information and services including high-resolution figures, can be found at: 

http://jn.physiology.org/cgi/content/full/82/6/2853 

Additional material and information about Journal of Neurophysiology can be found at: 

http://www.the-aps.org/publications/jn 

This information is current as of March 31, 2010 . 

Journal of Neurophysiology publishes original articles on the function of the nervous system. It is published 12 times a year 

(monthly) by the American Physiological Society, 9650 Rockville Pike, Bethesda MD 20814-3991. Copyright © 2005 by the 

American Physiological Society. ISSN: 0022-3077, ESSN: 1522-1598. Visit our website at http://www.the-aps.org/. 

Downloaded from 

jn.physiology.org 

on March 31, 2010

Inhibition of Rapid Heat Responses in Nociceptive Primary Sensory 

Neurons of Rats by Vanilloid Receptor Antagonists 

TIMO KIRSCHSTEIN, 1 WOLFGANG GREFFRATH, 1 DIETRICH BÜSSELBERG, 2 AND ROLF-DETLEF TREEDE 1 

1 Institute of Physiology and Pathophysiology, Johannes Gutenberg University, D-55099 Mainz; and 2 Center of Physiology 

and Pathophysiology, Georg August University, D-37073 Göttingen, Germany 

Kirschstein, Timo, Wolfgang Greffrath, Dietrich Büsselberg, and 

Rolf-Detlef Treede. Inhibition of rapid heat responses in nociceptive 

primary sensory neurons of rats by vanilloid receptor antagonists. J. 

Neurophysiol. 82: 2853–2860, 1999. Recent studies demonstrated that 

heat-sensitive nociceptive primary sensory neurons respond to the 

vanilloid receptor (VR) agonist capsaicin, and the first cloned VR is 

a heat-sensitive ion channel. Therefore we studied to what extent 

heat-evoked currents in nociceptive dorsal root ganglion (DRG) neurons 

can be attributed to the activation of native vanilloid receptors. 

Heat-evoked currents were investigated in 89 neurons acutely dissociated 

from adult rat DRGs as models for their own terminals using 

the whole cell patch-clamp technique. Locally applied heated extracellular 

solution (effective temperature �53°C) rapidly activated reversible 

and reproducible inward currents in 80% (62/80) of small 

neurons (�32.5 �m), but in none of nine large neurons (P � 0.001, 

� 2 test). Heat and capsaicin sensitivity were significantly coexpressed 

in this subpopulation of small DRG neurons (P � 0.001, � 2 test). 

Heat-evoked currents were accompanied by an increase of membrane 

conductance (320 � 115%; mean � SE, n � 7), had a reversal 

potential of 5 � 2mV(n � 5), which did not differ from that of 

capsaicin-induced currents in the same neurons (4 � 3 mV), and were 

carried at least by Na � and Ca 2� (pCa 2� � pNa � ). These observations 

are consistent with the opening of temperature-operated nonselective 

cation channels. The duration of action potentials was significantly 

higher in heat-sensitive (10–90% decay time: 4.45 � 0.39 ms, 

n � 12) compared with heat-insensitive neurons (2.18 � 0.19 ms, n � 

6; P � 0.005, Student’s t-test), due to an inflection in the repolarizing 

phase. This property as well as capsaicin sensitivity and small cell size 

are characteristics of nociceptive DRG neurons. When coadministered 

with heat stimuli, the competitive VR antagonist capsazepine (1 �M 

to 1 mM) significantly reduced heat-evoked currents in a dose-dependent 

manner (IC 50 13 �M, Hill slope �0.58, maximum effect 75%). 

Preincubation for 12–15 s shifted the IC 50 by �0.5 log 10 units to an 

estimated IC 50 of �4 �M. The noncompetitive VR antagonist ruthenium 

red (5 �M) significantly reduced heat-evoked currents by 33 � 

6%. The effects of both VR antagonists were rapidly reversible. Our 

results provide evidence for a specific activation of native VRs in 

nociceptive primary sensory neurons by noxious heat. The major 

proportion of the rapid heat-evoked currents can be attributed to the 

activation of these temperature-operated channels, and noxious heat 

may be the signal detected by VRs under physiological conditions. 

INTRODUCTION 

Polymodal C- and A�-fiber nociceptors in the skin of primates 

and humans respond within a few milliseconds to fast 

temperature increases generated by infrared lasers (Bromm and 

The costs of publication of this article were defrayed in part by the payment 

of page charges. The article must therefore be hereby marked “advertisement” 

in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 

0022-3077/99 $5.00 Copyright © 1999 The American Physiological Society 

Treede 1983; Tillman et al. 1995). These short latencies observed 

in vivo suggest a rapid direct transduction mechanism 

for heat stimuli at the peripheral endings of small dorsal root 

ganglion (DRG) neurons (Treede et al. 1995, 1998). DRG 

neurons are primary sensory neurons comparable to olfactory 

or visual receptor cells, but in contrast to vision and olfaction, 

little is known about the transduction mechanisms of nociception 

and pain (Belmonte 1996). 

Recent in vitro studies of the underlying membrane currents 

have supported the existence of a rapid heat transduction 

mechanism in nociceptive primary sensory neurons of the rat 

(Cesare and McNaughton 1996; Dittert et al. 1998; Kirschstein 

et al. 1997; Nagy and Rang 1999). The somata of DRG neurons 

are used as models for their own peripheral terminals 

(Vyklicky and Knotková-Urbancová 1996) because the terminals 

in the skin are inaccessible for adequate electrophysiological 

studies (patch-clamp) due to their small size and tough 

surrounding tissue. Brief heat stimuli of �1 s duration were 

found to elicit inward currents in DRG neurons of neonatal rats 

after 4–6 days in primary culture (Cesare and McNaughton 

1996) and within 4–30 h after acute dissociation from adult 

rats (Kirschstein et al. 1997). The activation was rapid but not 

instantaneous with a half-time of maximal activation of 35 ms 

and with a threshold temperature of �43°C (Cesare and Mc- 

Naughton 1996). These findings suggest a novel mechanism of 

rapid cellular signaling by temperature-operated ion channels. 

We have previously demonstrated that heat sensitivity in 

acutely dissociated DRG neurons is coexpressed with sensitivity 

to capsaicin (Kirschstein et al. 1997), the active ingredient 

in hot chili peppers that is a selective activator of nociceptive 

afferents (Szallasi and Blumberg 1996). This observation 

would be compatible with two possibilities: 1) capsaicin and 

heat act on different transduction pathways that are co-localized 

in the same nociceptive neurons, or 2) capsaicin and heat 

act on a common transduction pathway. The second hypothesis 

is more likely because the first cloned vanilloid receptor (VR1) 

is a nonselective cation channel that is activated by the vanilloid 

receptor agonist capsaicin and by raising the temperature 

to 40–45°C (Caterina et al. 1997; Tominaga et al. 1998). 

On the other hand, additional heat-transduction mechanisms 

have been described, which differ from the rapid transduction 

pathway with respect to 1) threshold temperature and response 

latency (Treede et al. 1995, 1998), 2) correlation with VR 

expression (Nagy and Rang 1999), and 3) dependence on 

intracellular calcium (Reichling and Levine 1997). 

The aim of this study was to test whether and to what extent 

heat-evoked inward currents in nociceptive DRG neurons can 

2853 



on March 31, 2010

2854 KIRSCHSTEIN, GREFFRATH, BÜSSELBERG, AND TREEDE 

be attributed to the activation of native vanilloid receptors. For 

this purpose, we characterized the electrophysiological properties 

of heat-evoked inward currents and of the neurons that 

express these currents, tested responses of the neurons to the 

VR-agonist capsaicin, and determined the effects of the two 

known vanilloid receptor antagonists, ruthenium red (RR) 

(Amann and Maggi 1991) and capsazepine (CPZ) (Bevan et al. 

1992), on rapid heat-evoked currents in acutely dissociated 

DRG neurons of adult rats. Preliminary accounts of this study 

have appeared in abstract form (Kirschstein et al. 1998). 

This paper contains essential parts of the dissertation of T. 

Kirschstein. 

METHODS 

The preparation, neuron dissociation, electrophysiological recordings, 

and heat stimulation were done as previously described (Kirschstein 

et al. 1997). Briefly, adult Sprague-Dawley rats (120–270 g) of 

both sexes were deeply anesthetized with diethylether and rapidly 

decapitated (see Greffrath et al. 1998 for detail). This method is in 

accordance with German national law and the rules of local ethical 

committees. The spine was chilled at 4°C in F12-Dulbecco’s modified 

Eagle’s medium (Sigma) saturated with carbogen gas (95% O 2-5% 

CO 2) and additionally containing 30 mM NaHCO 3 (Merck, Darmstadt, 

Germany), 100,000 units l �1 penicillin and 100 mg l �1 streptomycin 

(Sigma). Thoracic and lumbar DRGs (8–15) were quickly 

dissected and freed from connective tissue. Neurons were dissociated 

in an incubation chamber enriched with carbogen gas at 37°C using 

collagenase CLS II (5–10 mg ml �1 , 40–50 min; Biochrome, Berlin, 

Germany) and trypsine (0.2–1 mg ml �1 , 10–12 min; Sigma) dissolved 

in the F12 medium. After trituration (4–6 times with a Pasteur 

pipette) neurons were plated on 35-mm-diam culture dishes, which 

also served as recording chambers, and stored at 37°C in a humidified 

5% CO 2 atmosphere before being used for electrophysiological recordings 

3–12 h (up to 30 h in some cases) after dissociation. 

Electrophysiology 

Only round or oval-shaped neurons without any processes were 

included in this study. The average of the major and the minor 

diameter was used to measure the size of oval shaped neurons. Whole 

cell patch-clamp experiments were performed in carbogen gas saturated 

F12 medium (pH 7.4) at room temperature (RT) using an 

Axopatch 200A amplifier (Axon Instruments) in voltage-clamp mode 

at a holding potential of �80 mV controlled by pCLAMP6 software. 

Data were also registered on a chart recorder (L6512, Linseis, Selb, 

Germany). Patch pipettes were fabricated from borosilicate glass 

capillaries (Hilgenberg, Malsfeld, Germany) using a horizontal micropipette 

puller (P-87, Sutter) and filled with a solution containing (in 

mM) 160 KCl, 8.13 EGTA, and 10 HEPES (pH 7.2; R Tip � 5.3 � 0.2 

M�, mean � SE). Cell diameter, cross-sectional area, and membrane 

capacitance were measured, and excitability was tested by depolarizing 

voltage steps for each neuron. Cells lacking a fast inward current 

with a reversal potential close to the equilibrium potential of sodium 

followed by a prolonged outward current were excluded from further 

investigation. Experiments in current-clamp mode were performed to 

measure the resting membrane potential (RMP) of each neuron and to 

investigate single action potentials (APs) elicited by short (3 ms) 

depolarizing current pulses in neurons that were hyperpolarized by 

constant current injection resulting in membrane potentials between 

�70 and �80 mV (n � 18). Inflections in the repolarizing phase were 

qualitatively detected as second negative peak in the first derivative 

(dV/dt) of each AP, the duration of repolarization was quantitatively 

assessed by the 10–90% decay time. 

Heat stimulation, drug application, and solutions 

Application of �50 �l of heated extracellular solution through a 

puffing system fixed on a micromanipulator was used to elicit heatevoked 

currents. Control measurements with a fast temperature sensor 

(BAT-12, Physitemp; � � 5 ms) in place of the neurons revealed an 

effective peak temperature of �53°C, a rise time of �250 ms, and a 

decay with a time constant of �20 s. Effects were compared with 

those of application of the same amount of medium at RT. Heat 

stimuli with or without vanilloid receptor antagonists and control 

applications at RT were repeated 2–10 times, and the elicited currents 

were averaged. A neuron was considered heat sensitive when the 

heat-evoked inward current was significantly greater than any fluctuations 

caused by superfusion of solution at RT (unpaired 1-tailed 

Student’s t-test for each neuron, P � 0.05). Heating the buffered 

extracellular solution may change its pH, and acid solutions of pH 6.2 

are known to activate nociceptive DRG neurons (Bevan and Yeats 

1991). The pH of a HEPES-buffered solution decreases while heating 

(e.g., pH 7.1 at 50°C according to the Henderson-Hasselbalch equation). 

In contrast higher temperatures increase the pH of a NaHCO 3/ 

CO 2 buffer, because the solubility of CO 2 is reduced and thus reverses 

the HEPES effect. Off-line measurements showed that the pH of the 

F12 medium maximally changed in a range of 7.28–7.52 while 

heating to 50°C and cooling down to RT. Furthermore, the absence of 

color changes of the pH indicator phenol red, which was present in the 

extracellular solution, verified on-line that pH throughout all experiments 

remained well above 6.8. The membrane conductance was 

measured in voltage-clamp mode by hyperpolarizing pulses (5 mV, 10 

ms, 50 s �1 ), and conductance changes were determined at the maximum 

amplitude of heat evoked currents. 

Reversal potentials of heat- and capsaicin-induced currents were 

measured as described by Liu et al. (1997) using fast depolarizing 

ramps (�80 to �30 mV in 200 ms every 550 ms). In these experiments 

patch pipettes were filled with a potassium-free solution containing 

(in mM) 140 CsCl, 10 HEPES, 10 EGTA, and 4 MgCl 2 

(adjusted to pH 7.2). Tetrodotoxin (TTX, 100 �M; Sigma) and nifedipine 

(1 �M; Sigma) were added to the extracellular solution to block 

voltage-gated Na � and Ca 2� channels. These neurons were not included 

in the general statistics of heat-sensitive cells. To examine 

charge carriers of heat-induced currents, Na � in the extracellular 

solution was replaced by N-methyl-D-glucamine (NMDG; Sigma). In 

some of these experiments extracellular Ca 2� was raised to 10 mM. 

Reversibility of ion replacement experiments was tested by final 

application of heated solution at physiological ion composition. Capsaicin 

(Sigma) was initially dissolved in ethanol, diluted to its final 

concentration in F12 medium, and applied through the puffing system 

as described above. CPZ (dissolved in DMSO; purchased from RBI, 

Cologne, Germany) and RR (in extracellular solution; RBI) were 

prepared as concentrated stock solutions, diluted to final concentration 

in F12 medium, and applied either at RT or �53°C in the same way. 

Because CPZ solutions contained final concentrations of 0.01–10% 

DMSO, the same amount was added to all solutions applied in CPZ 

experiments, too. In the concentration range of 0.01–1%, DMSO 

alone did not elicit any visible effects, but with 10% DMSO the 

proportion of neurons that could not be kept for the full protocol was 

increased. Reversibility of antagonist action was tested by reapplication 

of heated extracellular solution without any agents (n � 21). Only 

neurons tested for reversibility were included for further statistical 

analysis. 

Data analysis 

Off-line measurements and statistical analysis were done using 

pCLAMP6 (Axon Instruments) and EXCEL 5.0 (Microsoft). Data are 

presented as means � SE. Treatment effects were statistically analyzed 

by Student’s t-test for paired data and � 2 test for analysis of 

incidences. Dose dependence of the CPZ-induced inhibition was 



on March 31, 2010

analyzed by a repeated measures ANOVA with Student-Newmann- 

Keuls post hoc test for ordered means (STATISTICA 4.5, StatSoft). 

Error probabilities P � 0.05 were considered statistically significant. 

Fitting of the dose-response function was done using GraphPad Prism 

2.01 (GraphPad Software). 

RESULTS 

Electrophysiological properties of heat-sensitive 

DRG neurons 

Whole cell patch-clamp experiments were performed on 89 

acutely dissociated DRG neurons clamped at �80 mV. Brief 

superfusion of heated extracellular solution (effective temperature 

�53°C) activated a significantly greater inward current 

than the same amount of solution applied at RT (see METHODS) 

in 62 of the 80 small neurons (80%; see Fig. 1A for an 

example). In contrast, none of nine large neurons (diameter 

�32.5 �m) significantly responded to heat (P � 0.001, � 2 

test). In the majority of heat-sensitive neurons examined (47 of 

62) application of solution at RT did not elicit any change in 

holding currents. Currents elicited by control applications at 

RT were even significantly smaller in heat-sensitive (10 � 3 

pA, n � 62) than in heat-insensitive neurons (53 � 17 pA, n � 

18, P � 0.001). Within heat-insensitive neurons, however, the 

inward currents elicited by solutions at either �53°C (62 � 16 

pA) or RT (53 � 17 pA) did not differ significantly. Heatevoked 

currents were reproducible at 1 to 15 s interstimulus 

intervals (1st: 279 � 43 pA; 2nd: 310 � 44 pA, n � 62, n.s.) 

and were accompanied by an increase in membrane conductance 

(Fig. 1B), consistent with the opening of temperatureoperated 

channels. The averaged maximum increase of the 

membrane conductance during the peak of heat-evoked currents 

was 320 � 115% (n � 7). 

Within this subpopulation of small DRG neurons (diameter 

FIG. 1. Heat-evoked inward currents are accompanied by an increase of 

membrane conductance. A: superfusion of heated extracellular solution 

(�53°C) caused a large inward current in a small dorsal root ganglion (DRG) 

neuron [diameter 22.5 �m, resting membrane potential (RMP) ��74 mV, 

V Hold ��80 mV]. The increase of membrane conductance was measured by 

repetitive application of hyperpolarizing voltage pulses (5 mV, 10 ms, 50 s �1 ) 

throughout the experiment. Interruption of the trace of �1 s is due to the 

acquisition mode. B: the amplification (5-fold) of the current trace above as 

indicated by rectangular boxes demonstrates the fully reversible increase of 

membrane conductance. 

NOCICEPTOR HEAT RESPONSES AND VANILLOID RECEPTORS 

2855 

FIG. 2. Heat-sensitive and heat-insensitive small DRG neurons differ with 

respect to action potential shape. Different shapes of action potentials (APs) 

elicited by depolarizing pulses (indicated by solid lines) in a heat-sensitive (A) 

and a heat-insensitive (B) small DRG neuron (A � 27.5 �m, both). APs are 

averages of 20 single pulses, each. A: the heat-sensitive neuron (RMP ��43 

mV, hyperpolarized to �70 mV by constant current injection) displayed an AP 

with a prominent shoulder in the repolarizing phase and a duration of 4.4 ms 

(width at half-maximum). B: AP of a heat-insensitive neuron (RMP ��49 

mV, hyperpolarized to �78 mV) without a pronounced inflection and a 

duration of 2.6 ms measured at half-maximum amplitude. C: repolarization 

measured as 10–90% decay time in heat-sensitive (●) and heat-insensitive (E) 

neurons; means are indicated by dotted lines. A prominent shoulder in the AP 

repolarizing phase was seen in all neurons with a decay time of approximately 

�2.7 ms (dashed line). This shoulder was significantly associated with heat 

sensitivity of the neurons (P � 0.005, � 2 test). 

�32.5 �m), heat-sensitive and heat-insensitive neurons did not 

differ in diameter (27.5 � 0.3 �m for heat-sensitive, 27.4 � 

0.8 �m for heat-insensitive neurons) nor in whole cell capacitance 

(23.2 � 0.9 pF vs. 21.2 � 1.7 pF). RMP was more 

depolarized in heat-sensitive (�46 � 1 mV) than in heatinsensitive 

small neurons (�52 � 2 mV, P � 0.033), but there 

was no correlation between RMP, measured in current-clamp 

mode, and heat-evoked current in neurons clamped at �80 mV 

in voltage-clamp mode (r ��0.17, n.s., n � 62). Moreover, 

the three cells with the most negative RMP of about �70 mV 

exhibited three of the largest heat responses (�600 pA). Therefore 

heat sensitivity is not a function of a depolarized membrane 

potential. 

A prominent inflection in the repolarizing phase of APs 



on March 31, 2010


FIG. 3. Capsazepine reduces rapid heat-evoked inward currents reversibly and dose dependently. Effect of the competitive 

vanilloid receptor antagonist capsazepine (CPZ) on the heat-activated inward current. A: superfusion with heated extracellular 

solution at �53°C elicited an inward current of 240 pA in a heat-sensitive neuron (A � 30 �m, RMP ��47 mV, V Hold ��80 

mV). B: coapplication of CPZ (10 �M) and heat reduced the heat-induced current to 120 pA. C: when heated control solution was 

applied again, the current recovered to 220 pA. D: averaged heat-evoked inward currents. The current was initially 231 � 35 pA. 

CPZ (10 �M) significantly reduced the current to 150 � 20 pA (**P � 0.01, Student’s paired t-test). When tested �60 s later with 

control solution the heat-evoked current was again 228 � 31 pA. E: dose-dependent CPZ-induced inhibition of normalized 

heat-evoked inward currents (divided by the average of before and after the CPZ application). CPZ reduced the heat-evoked inward 

current to 85 � 6% at 1 �M, and significantly to 69 � 5% at 10 �M, to 42 � 2% at 100 �M, and to 31 � 9% at 1 mM (**P � 

0.01, ***P � 0.001, ANOVA with Student-Newmann-Keuls post hoc test). 

evoked by constant current injection was observed in 11 of 12 

heat-sensitive neurons (Fig. 2), whereas a smaller inflection 

was detectable only in 1 of 6 heat-insensitive small neurons 

(P � 0.005, � 2 test). As a consequence of this “shoulder,” 

action potentials in heat-sensitive neurons displayed significantly 

longer repolarizing phases (10–90% decay time: 4.45 � 

0.39 ms, n � 12) than heat-insensitive neurons (2.18 � 0.19 

ms, n � 6; P � 0.005). 

To test whether heat and capsaicin sensitivity are coexpressed 

in DRG neurons, the effect of capsaicin (1–10 �M) 

was examined in 29 neurons. None of 5 large neurons (diameter 

�32.5 �m) but 16 of 24 small neurons (�32.5 �m) were 

excited by capsaicin (P � 0.01, � 2 test). Only 1 of 15 heatsensitive 

neurons was not excited by capsaicin, whereas 12 of 

14 heat-insensitive neurons were also capsaicin insensitive 

(P � 0.001, � 2 test). Two heat-insensitive neurons were ex- 

FIG. 4. Ruthenium red reversibly reduces 

heat-evoked inward currents. Effect of the 

noncompetitive vanilloid receptor antagonist 

ruthenium red (RR) on the heat-activated inward 

current. A: superfusion with heated extracellular 

solution at 53°C elicited an inward 

current of 360 pA in a heat-sensitive neuron 

(A � 25 �m, RMP ��52 mV, V Hold ��80 

mV). B: coapplication of RR (5 �M) and heat 

reduced the heat-induced current to 160 pA. 

C: when heated control solution was applied 

again, the current recovered to 340 pA. D: 

averaged heat-evoked inward currents. The 

current was initially 231 � 35 pA. RR (5 

�M) significantly reduced the current to 

161 � 16 pA (P � 0.05, Student’s paired 

t-test). When tested �60 s later with control 

solution, the heat-evoked current was again 

252 � 47 pA. E: normalized heat-evoked 

inward currents (as in Fig. 3E). RR reduced 

the heat-evoked inward current to 67 � 6% 

(P � 0.005). 



on March 31, 2010

cited by capsaicin, but one of those neurons developed heat 

sensitivity after the capsaicin-induced excitation, the remaining 

neuron was not tested with heat after capsaicin. Time-to-peak 

of capsaicin-induced currents varied between 300 ms and 65 s 

with a mean of 14 � 5s(n � 14). Capsaicin often induced 

multiple current peaks, and for the value above the time to the 

maximum current was measured (2,950 � 700 pA). Whereas 

some capsaicin-induced currents were almost as fast as heatevoked 

currents, others were as slow as the long-latency capsaicin 

currents reported by others (Liu and Simon 1996; Petersen 

et al. 1996). 

The reversal potentials of heat-evoked currents and capsaicin-induced 

currents were determined in five small neurons 

(diameter 28 � 0.8 �m) with current-voltage (I-V) curves 

elicited by fast depolarizing ramps using Cs � in the intra- and 

TTX (100 �M) and nifedipine (1 �M) in the extracellular 

solution. These agents did not abolish all inward currents 

elicited by the initial depolarizing ramp, which may be due to 

the presence of, e.g., TTX-resistant sodium channels. However, 

these small currents disappeared during repeated ramps. 

Therefore the repeated-ramps protocol was run until those 

currents had inactivated (within �100 cycles), before heat or 

capsaicin were applied to determine reversal potentials. The 

reversal potential of heat-evoked currents was 5 � 2 mV, that 

of the initial phase of capsaicin-induced currents (1 �M) in the 

same neurons 4 � 3 mV (n.s.). Within the variability between 

neurons, the reversal potentials of heat-evoked and capsaicininduced 

currents were highly correlated (r � 0.93, P � 0.05, 

n � 5). The reversal potential near 0 mV can be explained by 

a nonselective cation channel, that is outwardly permeable to 

Cs � . In conclusion, heat-evoked currents and capsaicin-induced 

currents are both carried through similar nonspecific 

cation channels. 

We therefore tested whether the rapid heat-induced inward 

currents are carried by Na � and/or Ca 2� . When Na � was 

replaced by NMDG (n � 15), heat-evoked currents were 

reversibly reduced to 34 � 4% of the mean currents before and 

after ion replacement (P � 0.001). When Ca 2� in the Na � -free 

extracellular solution was raised to 10 mM (n � 7), heatevoked 

currents were enhanced to 478 � 120% (P � 0.05). 

Although in these experiments the proportion of putative 

charge carriers was smaller than in physiological extracellular 

solution (10 vs. 145 mM) heat-evoked currents did not differ 

significantly from the normalized controls (129 � 38% vs. 

100%; n.s.), indicating a higher permeability for Ca 2� than for 

Na � ions. 

Effects of vanilloid receptor antagonists on heat-evoked 

currents 

Heat-sensitive neurons were tested for the effects of vanilloid 

receptor antagonists on heat-evoked inward currents to 

examine pharmacologically, whether and to what extent native 

vanilloid receptors contribute to rapid heat-transduction mechanisms. 

When the competitive antagonist CPZ (10 �M) was 

added to the heated extracellular solution (Fig. 3, A–C) heatevoked 

inward currents were significantly reduced by 31 � 5% 

(P � 0.01, n � 12). This inhibition was rapidly and fully 

reversible, as demonstrated by the response to the application 

of heated control solution �60 s after the CPZ application, 

which did not differ from the initial heat response (Fig. 3D). 


Furthermore, CPZ significantly increased the time-to-peak of 

heat-evoked currents from 124 � 5msto154� 10 ms (P � 

0.01; n � 12). In contrast, in normal extracellular solution, 

smaller currents were associated with shorter time-to-peak (r � 

0.64, P � 0.05, n � 12). 

The CPZ-induced inhibition of heat-evoked currents was 

dose dependent (1 �M to1mM;F 4,38 � 6.42, P � 0.001, 

ANOVA; see Fig. 3E) with an IC 50 of 13 �M (log 10 IC 50 � 

�4.88 � 0.18), a Hill slope of �0.58 � 0.10, and an extrapolated 

efficacy of 75 � 5%, suggesting that most of the rapid 

heat-evoked current was inhibited by CPZ, although CPZ 

binding probably was not at equilibrium with simultaneous 

application. When neurons were preincubated with 10 �M 

CPZ (i.e., at the steepest part of the dose-response function) for 

12–15 s, heat-evoked currents were reversibly reduced by 

47.2 � 8% of the normalized controls (P � 0.001; n � 7). The 

difference between the inhibiting effect of CPZ with and without 

preincubation was marginally significant (47 vs. 31%; P � 

0.11), indicating a very fast action of the competitive antagonist 

on heat-evoked currents. This difference corresponded to a 

shift of the dose-response curve by �0.5 log 10 units resulting 

in an estimated IC 50 of �4 �M. Application of CPZ at room 

temperature did not elicit any currents (0 � 0 pA, n � 2, data 

not shown). 

Application of RR (5 �M) significantly reduced heat responses 

by 33 � 6% (Fig. 4; P � 0.005, n � 7), likewise in a 

fully reversible manner and without affecting neurons when 

applied at RT (0 � 0 pA, n � 3, data not shown). RR 

specifically inhibits the vanilloid receptors at lower concentrations 

only in a narrow range (estimated to 0.1–10 �M) and may 

also reduce intracellular calcium release (Amann and Maggi 

1991; Maggi 1991). Thus we did not use higher concentrations. 

DISCUSSION 

2857 

A subpopulation of small neurons acutely dissociated from 

rat dorsal root ganglia responded to brief noxious heat stimuli 

with inward currents. In contrast to heat-insensitive small and 

large neurons, this population was excited by the vanilloid 

receptor agonist capsaicin and exhibited a prolonged action 

potential duration with a prominent inflection in the repolarization 

phase. Heat-sensitive DRG neurons therefore fulfilled 

several criteria for the somata of primary nociceptive afferents 

(Gold et al. 1996; Harper and Lawson 1985; Petersen and 

LaMotte 1991). Heat-evoked currents were accompanied by a 

conduction increase, had a reversal potential near 0 mV, and 

were permeable for Ca 2� � Na � . These findings suggest that 

the currents were carried through temperature-operated nonspecific 

cation channels. The reversible inhibition of heatevoked 

inward currents by the competitive VR antagonist CPZ 

and the noncompetitive VR antagonist RR implies the involvement 

of vanilloid receptors in this rapid transduction pathway 

for noxious heat. 

Properties of heat-sensitive DRG neurons 

As in our previous study (Kirschstein et al. 1997; see also 

Dittert et al. 1998), we found rapid heat-evoked currents only 

in small neurons (�32.5 �m). Nociceptive neurons are generally 

small, consistent with their small axons (Gold et al. 1996; 

Harper and Lawson 1985; Petersen and LaMotte 1991). In their 



on March 31, 2010


pioneering study on heat-evoked currents, Cesare and Mc- 

Naughton (1996) therefore restricted their sample to small 

DRG neurons. Two other studies reported heat-evoked currents 

in some larger DRG neurons (Nagy and Rang 1999; Reichling 

and Levine 1997), but these responses are likely due to other 

heat transduction mechanisms (see Multiple heat transduction 

pathways). 

Rapid heat-evoked inward currents in DRG neurons were 

associated with comparably fast responses to the vanilloid 

receptor agonist capsaicin, confirming the results of our previous 

study (Kirschstein et al. 1997). A similar coexpression has 

been reported in rat and monkey nociceptive afferents in vivo 

(Baumann et al. 1991; Szolcsányi et al. 1988). The finding that 

vanilloid receptor antagonists substantially reduced all rapid 

heat responses in the present study provides another line of 

evidence that heat-sensitive neurons express vanilloid receptors. 

The repolarization phase of the AP was significantly longer 

in heat-sensitive than heat-insensitive small DRG neurons, due 

to a shoulder that was exhibited by 11 of 12 heat-sensitive but 

only 1 of 6 heat-insensitive neurons. This AP shape has been 

associated with small soma diameter and slow axonal conduction 

velocity in the C-fiber range (Harper and Lawson 1985), 

the presence of TTX-resistent sodium channels (Waddell and 

Lawson 1990), as well as responsiveness to the vanilloid 

receptor agonist capsaicin (Del Mar et al. 1996). Our data 

support the view that somal AP shape is associated with 

functional characteristics of the cell as a nociceptive primary 

sensory neuron rather than simply with its size. 

Properties of rapid heat-evoked currents 

The suggestion that nociceptive primary sensory neurons 

possess a rapid transduction mechanism for noxious heat stimuli 

that is independent from tissue damage (Treede et al. 1995) 

was supported by the demonstration that noxious heat elicited 

inward currents in small DRG neurons (Cesare and McNaughton 

1996). Two possible mechanisms may be responsible for 

the inward currents: 1) the heat-induced opening of membrane 

channels or 2) the heat-induced inactivation of persistent outward 

currents. Because in our data the heat-evoked inward 

currents were accompanied by significant increases in whole 

cell conductance, we conclude that they were due to channel 

opening. Likewise, a preliminary report on cell-attached patch 

recordings has demonstrated channel opening by heat stimuli 

of 40–48°C (Nagy and Rang 1998). 

Noxious heat may either induce a direct opening of temperature-operated 

channels or trigger an intracellular signal cascade 

resulting in channel opening. A direct transduction mechanism 

seems more probable, because the time-to-peak of heatevoked 

currents was at least as fast as the temperature peak of 

the stimulus used. This finding is consistent with observations 

on primary nociceptive afferents in vivo, that the initial burst of 

action potentials is already generated during the rise time of the 

cutaneous heat stimulus (Treede et al. 1995). To determine the 

activation kinetics exactly, a heat stimulus has to be established, 

which is much faster than the rapid heat-evoked currents 

(e.g., a laser-stimulator described by Baumann and Martenson 

1994). 

The reversal potential of heat-evoked currents in our study 

was similar to that in cultured DRG neurons of neonatal rats 

(Cesare and McNaughton 1996), which resembled that of the 

rapid component of capsaicin-induced currents (Liu and Simon 

1996; Oh et al. 1996). Ion replacement experiments showed 

that the temperature-operated channels are permeable to Na � , 

Ca 2� , and Cs � (cf. Cesare and McNaughton 1996). Finally, 

rapid heat-evoked currents had a higher permeability for Ca 2� 

than for Na � ions, which is also typical for capsaicin-induced 

currents (Bevan and Szolcsányi 1990; Jung et al. 1999). From 

changes of reversal potentials of heat-evoked currents after ion 

replacement, the permeability ratio pNa � :pCa 2� was calculated 

to be 1:1.28 (Cesare and McNaughton 1996). In addition 

to these similarities of heat-evoked and capsain-induced currents 

described in different studies, we now have demonstrated 

that the reversal potential of both currents is not distinguishable 

in the same neuron. Thus both currents may be conducted at 

least in part by identical channels. 

Vanilloid receptors and heat transduction 

A candidate for such a channel is the vanilloid receptor VR1, 

a nonselective cation channel that is activated by heating the 

cell to 40–45°C and that is selectively expressed in DRGs and 

trigeminal ganglia (Caterina et al. 1997; Tominaga et al. 1998). 

The heterologous expression of VR1 in both human embryonic 

kidney cells (HEK 293) and frog oocytes conveyed heat sensitivity 

to these cells (Caterina et al. 1997), consistent with a 

direct action of noxious heat on capsaicin-sensitive membrane 

channels. Single-channel openings were observed in excised 

patches of these VR1-transfected cells to both heat and capsaicin 

(Tominaga et al. 1998). In the present study heat-evoked 

currents in DRG neurons were inhibited by 75% by the competitive 

VR antagonist CPZ and by 33% by the noncompetitive 

VR antagonist RR. Thus we have now provided pharmacological 

evidence that native vanilloid receptors are a key element 

in the responses of nociceptive primary sensory neurons to 

noxious heat stimuli. 

The reduction in heat-evoked currents by 10 �M CPZ (by 

31% with co-application and 47% with preincubation) was 

smaller than that reported for currents elicited by 0.5 �M 

capsaicin (by 96%) in neonatal rat DRG neurons (Bevan et al. 

1992). Whereas the reduction of heat-evoked currents by CPZ 

was fully reversible on wash out, the capsaicin-evoked currents 

recovered only by �50%; this was attributed to desensitization 

induced by the first application of capsaicin (Bevan et al. 

1992). Interestingly, in Xenopus oocytes expressing the cloned 

VR1, 10 �M CPZ had 70% efficacy against 44°C (Tominaga 

et al. 1998), but �95% efficacy against 0.6 �M capsaicin 

(Caterina et al. 1997). In HEK 293 cells, CPZ had 90% efficacy 

against moderate noxious heat and 97% against capsaicin, but 

the preincubation with CPZ was four times longer for the heat 

tests than for capsaicin, recovery was incomplete, and repetition 

of the 46°C heat stimulus by itself reduced the response by 

44% (Tominaga et al. 1998). Thus a reduced efficacy of CPZ 

to inhibit VR1 activation by heat compared with capsaicin is 

supported by these findings, too. Because protein structures are 

highly sensitive to temperature, the affinity of vanilloid receptors 

for CPZ is likely to be reduced at the temperature used in 

the present study. For enzymes such as lactate dehydrogenase, 

the Km for substrate binding is about three to five times higher 

at 50°C than at room temperature (Somero 1995). The temperature 

dependence of ligand-receptor interactions may explain 



on March 31, 2010

most of the difference between the IC 50 described here at 

�53°C (4–13 �M) and the binding constant of CPZ to vanilloid 

receptors at room temperature (K d of 0.1–0.7 �M), that 

was estimated from Schild plots of the competitive antagonism 

of CPZ versus capsaicin and resiniferatoxin in rat DRG neurons 

(Bevan et al. 1992). Unfortunately, a Schild analysis to 

determine the K d for CPZ-induced inhibition of heat-evoked 

currents is impracticable, because the upper end of useful heat 

intensities (i.e., the “agonists” concentration) is limited by 

activation of another transduction mechanism that is VR independent 

(Caterina et al. 1999; Nagy and Rang 1999) (see 

Multiple heat transduction pathways) and ultimately by cell 

damage. Therefore we did not determine, whether CPZ inhibits 

heat-evoked currents competitively like it antagonizes capsaicin-induced 

currents. 

Alternatively, the apparent difference in affinity raises the 

question, whether the inhibition of heat-evoked currents may 

have been unrelated to a specific inhibition of vanilloid receptors. 

CPZ had no effects on GABA-receptors, ATP-receptors, 

and depolarization-induced ion fluxes at RT (Bevan et al. 

1992). Recently, however, nonspecific effects of CPZ have 

been described on voltage-activated calcium channels with an 

ED 50 � 8 �M (Docherty et al. 1997) and on nicotinic acetylcholine 

receptors at 10 �M (Liu and Simon 1997). The time to 

reach half-maximum effect on voltage-activated calcium channels 

was �5 min at 1–10 �M and 1 min at 100 �M, and this 

effect was irreversible. Likewise, the inhibition of nicotinic 

acetylcholine receptors by 10 �M CPZ was found with a 2-min 

preincubation period of the antagonist, and it took �10 min 

wash out to reverse this effect. Because in our data the antagonists 

were preincubated for a few seconds only and the 

inhibition of the “agonist” heat was fully reversible as soon as 

tested (�1 min or less), the inhibition of heat-evoked inward 

currents by CPZ is unlikely to be due to nonspecific effects. A 

further argument for the inhibition of heat-evoked currents by 

a specific interaction with vanilloid receptors derives from the 

significant effect of 5 �M RR. RR interacts with a different site 

of the vanilloid receptors (Bevan et al. 1992; Dray et al. 1990), 

and our dose was within the range that is considered specific 

for vanilloid receptors (0.1–10 �M) (Maggi 1991). 

Thus whereas nonspecific effects of CPZ have been reported, 

the temporal characteristics of those effects are sufficiently 

different from the rapid and fully reversible inhibition 

of heat-evoked inward currents by CPZ to warrant the conclusion 

that vanilloid receptors are essentially involved in the 

rapid heat transduction pathway. The receptor subtype need not 

necessarily be VR1, because multiple vanilloid receptors exist 

(Szallasi 1994), which may be coexpressed in the same neuron 

(Liu and Simon 1996; Liu et al. 1997; Petersen et al. 1996), and 

subtype specific antagonists are not available. 

Multiple heat transduction pathways 

The major proportion of the rapid heat-evoked current of the 

present study can be attributed to vanilloid receptor activation. 

However, the extrapolated efficacy of CPZ (75 � 5%) raises 

the question of whether this reflects a true incomplete antagonism 

(e.g., because heat acts at a different site of VR1 than 

CPZ) or whether the remaining fraction was due to another 

transduction mechanism that was independent of VRs. 

A subpopulation of cultured DRG neurons has recently been 


2859 

described, that were heat-sensitive but not excited by capsaicin 

(Nagy and Rang 1999), and a membrane channel with similar 

properties has been cloned and called vanilloid receptor–like 

channel (VRL-1) (Caterina et al. 1999). These neurons had a 

higher heat threshold, smaller maximum currents, and larger 

diameters. A corresponding heat transduction mechanism is 

present in a subclass of A�-fiber nociceptors that respond to 

heat only after prolonged stimulus duration or subsequent to 

tissue damage and are called HTM for high-threshold mechanoreceptor 

or type 1 AMH (the 1st type of A-fiber mechanoheat 

nociceptor discovered) (Fitzgerald and Lynn 1977; Perl 

1968; Treede et al. 1995). In an in vivo study on the rat 

saphenous nerve, all polymodal C-fibers and the majority of 

polymodal A�-fibers responded to close arterial injection of 

capsaicin, whereas the A�-fiber HTMs were not affected 

(Szolcsányi et al. 1988). Thus in type 1 AMH the slow heat 

transduction mechanism appears to be independent of vanilloid 

receptors and may be mediated by VRL-1. 

The remaining current after maximum blockade of VRs in 

the present study was significantly delayed, but the time-topeak 

was still an order of magnitude faster than that of currents 

presented by Nagy and Rang (1999) and Caterina et al. (1999). 

Moreover, the size distributions of neurons expressing VR1 

and VRL-1 do not suggest a widespread coexpression but an 

expression of either VR1 or VRL-1 (Caterina et al. 1999). In 

contrast to Nagy and Rang (1999) we found only one capsaicin-insensitive 

neuron that responded to heat. This difference 

between the studies may be due to several reasons. 1) The 

temperature used by us may not sufficiently activate the highthreshold 

current. 2) The stimulus used may have been too 

short; activation of the corresponding neurons in vivo (i.e., 

type 1 AMH) depends on a stimulus duration of several seconds 

(Treede et al. 1998). 3) Culture conditions were different 

(acutely dissociated vs. several days in culture with nerve 

growth factor, NGF); in the absence of NGF, type 1 AMHs in 

vivo change their phenotype into D-hair receptors (Ritter et al. 

1993). 

Still another mechanism seems to underly the heat-induced 

inward current (I heat) reported by Reichling and Levine (1997). 

Peak currents (50 vs. 280 pA) and conductance increases (25 

vs. 320%) were about one order of magnitude smaller than in 

the present and other studies (Cesare and McNaughton 1996; 

Kirschstein et al. 1997). Their I heat was blocked by extracellular 

cesium ions, whereas the nonspecific cation channel activated 

by brief noxious heat stimuli was even more permeable 

to Cs � than to Na � ions (Cesare and McNaughton 1996). 

Moreover, their I heat exhibited a linear increase from room 

temperature to 45°C. This temperature dependence is not easily 

compatible with nociceptive neurons. 

In summary, rapid heat responses in nociceptive primary 

sensory neurons appear to depend predominantly on a vanilloid 

receptor as the heat-sensing element. Conversely, our results 

support the hypothesis that the physiological function of the 

native vanilloid receptor VR1 is to detect noxious heat. This 

mechanism is different from two other heat transduction pathways 

that have been partly described, and the mechanisms of 

which await further studies. 

The authors thank G. Günther for help with the experiments, J. Rupp and H. 

Nawrath for the dose-response function analysis, W. Lang for evaluation of pH 



on March 31, 2010


changes in heated solutions, and G. Böhmer, P. Heusler, and W. Magerl for 

critical reading of the manuscript. 

This research was supported by the Deutsche Forschungsgemeinschaft 

(Tr236/11-1). 

Address for reprint requests: R.-D. Treede, Institute of Physiology and 

Pathophysiology, Johannes Gutenberg University, Saarstr. 21, D-55099 

Mainz, Germany. 

Received 21 January 1999; accepted in final form 28 July 1999. 

REFERENCES 

AMANN, R.AND MAGGI, C. A. Ruthenium red as a capsaicin antagonist. Life 

Sci. 49: 849–856, 1991. 

BAUMANN, T.K. AND MARTENSON, M. E. Thermosensitivity of cultured trigeminal 

neurons. Soc. Neurosci. Abstr. 20: 1379, 1994. 

BAUMANN, T. K., SIMONE, D. A., SHAIN, C.N.,AND LAMOTTE, R. H. Neurogenic 

hyperalgesia: the search for the primary cutaneous afferent fibers that 

contribute to capsaicin-induced pain and hyperalgesia. J. Neurophysiol. 66: 

212–227, 1991. 

BELMONTE, C. Signal transduction in nociceptors: general principles. In: Neurobiology 

of Nociceptors, edited by C. Belmonte and F. Cervero. New York: 

Oxford Univ. Press, 1996, p. 243–257. 

BEVAN, S., HOTHI, S., HUGHES, G., JAMES, I. F., RANG, H. P., SHAH, K., 

WALPOLE, C.S.J., AND YEATS, J. C. Capsazepine: a competitive antagonist of 

the sensory neurone excitant capsaicin. Br. J. Pharmacol. 107: 544–552, 

1992. 

BEVAN, S.AND SZOLCSÁNYI, J. Sensory neuron-specific actions of capsaicin: 

mechanisms and applications. Trends Pharmacol. Sci. 11: 330–333, 1990. 

BEVAN, S.AND YEATS, J. Protons activate a cation conductance in a subpopulation 

of rat dorsal root ganglion neurones. J. Physiol. (Lond.) 433: 145–161, 

1991. 

BROMM, B. AND TREEDE, R.-D. CO2 laser radiant heat pulses activate C 

nociceptors in man. Pflügers Arch. 399: 155–156, 1983. 

CATERINA, M. J., ROSEN, T. A., TOMINAGA, M., BRAKE, A.J.,AND JULIUS, D. 

A capsaicin-receptor homologue with a high threshold for noxious heat. 

Nature 398: 436–441, 1999. 

CATERINA, M. J., SCHUMACHER, M. A., TOMINAGA, M., ROSEN, T. A., LEVINE, 

J. D., AND JULIUS, D. The capsaicin receptor: a heat-activated ion channel in 

the pain pathway. Nature 389: 816–824, 1997. 

CESARE, P.AND MCNAUGHTON, P. A novel heat-activated current in nociceptive 

neurons and its sensitization by bradykinin. Proc. Natl. Acad. Sci. USA 

93: 15435–15439, 1996. 

DEL MAR, L. P., CARDENAS,C.G.,AND SCROGGS, R. S. Capsaicin preferentially 

affects small-diameter acutely isolated rat dorsal root ganglion cell bodies. 

Exp. Brain Res. 111: 30–34, 1996. 

DITTERT, I., VLACHOVÁ, V., KNOTKOVÁ, H., VITÁSKOVÁ, Z., VYKLICKY, L., 

KRESS, M., AND REEH, P. W. A technique for fast application of heated 

solutions of different composition to cultured neurones. J. Neurosci. Methods 

82: 195–201, 1998. 

DOCHERTY, R. J., YEATS, J. C., AND PIPER, A. S. Capsazepine block of 

voltage-activated calcium channels in adult rat dorsal root ganglion neurones 

in culture. Br. J. Pharmacol. 121: 1461–1467, 1997. 

DRAY, A., FORBES, C.A., AND BURGESS, G. M. Ruthenium red blocks the 

capsaicin-induced increase in intracellular calcium and activation of membrane 

currents in sensory neurones as well as the activation of peripheral 

nociceptors in vitro. Neurosci. Lett. 110: 52–59, 1990. 

FITZGERALD, M.AND LYNN, B. The sensitization of high threshold mechanoreceptors 

with myelinated axons by repeated heating. J. Physiol. (Lond.) 

365: 549–563, 1977. 

GOLD, M. S., DASTMALCHI, S., AND LEVINE, J. D. Co-expression of nociceptor 

properties in dorsal root ganglion neurons from the adult rat in vitro. 

Neuroscience 71: 265–275, 1996. 

GREFFRATH, W., MARTIN, E., REUSS, S., AND BOEHMER, G. Components of 

after-hyperpolarization in magnocellular neurones of the rat supraoptic 

nucleus in vitro. J. Physiol. (Lond.) 513: 493–506, 1998. 

HARPER, A.A. AND LAWSON, S. N. Electrical properties of rat dorsal root 

ganglion neurones with different peripheral nerve conduction velocities. 

J. Physiol. (Lond.) 359: 47–63, 1985. 

JUNG, J., HWANG, S. W., KWAK, J., LEE, S. Y., KANG, C. J., KIM, W. B., KIM, 

D., AND OH, U. Capsaicin binds to the intracellular domain of the capsaicinactivated 

ion channel. J. Neurosci. 19: 529–538, 1999. 

KIRSCHSTEIN, T., BÜSSELBERG, D., AND TREEDE, R.-D. Coexpression of heatevoked 

and capsaicin-evoked inward currents in acutely dissociated rat 

dorsal root ganglion neurons. Neurosci. Lett. 231: 33–36, 1997. 

KIRSCHSTEIN, T., BÜSSELBERG, D., AND TREEDE, R.-D. The vanilloid receptor 

antagonists ruthenium red and capsazepine reduce heat-evoked inward currents 

in rat DRG neurons. Soc. Neurosci. Abstr. 24: 384, 1998. 

LIU, L., LO, Y. C., CHEN,I.J.,AND SIMON, S. A. The responses of rat trigeminal 

ganglion neurons to capsaicin and two nonpungent vanilloid receptor agonists, 

olvanil and glyceryl nonamide. J. Neurosci. 17: 4101–4111, 1997. 

LIU, L.AND SIMON, S. A. Capsaicin-induced currents with distinct desensitization 

and Ca 2� dependence in rat trigeminal ganglion cells. J. Neurophysiol. 

75: 1503–1514, 1996. 

LIU, L.AND SIMON, S. A. Capsazepine, a vanilloid receptor antagonist, inhibits 

nicotinic acetylcholine receptors in rat trigeminal ganglia. Neurosci. Lett. 

228: 29–32, 1997. 

MAGGI, C. A. Capsaicin and primary afferent neurons: from basic science to 

human therapy? J. Auton. Nerv. Syst. 33: 1–14, 1991. 

NAGY, I.AND RANG, H. P. Noxious heat-activated microscopic currents in rat 

dorsal root ganglion neurones (Abstract). J. Physiol. (Lond.) 507: 29P, 1998. 

NAGY, I.AND RANG, H. Noxious heat activates all capsaicin-sensitive and also 

a sub-population of capsaicin-insensitive dorsal root ganglion neurons. 

Neuroscience 88: 995–997, 1999. 

OH, U., HWANG, S.W., AND KIM, D. H. Capsaicin activates a nonselective 

cation channel in cultured neonatal rat dorsal root ganglion neurons. J. Neurosci. 

16: 1659–1667, 1996. 

PERL, E. R. Myelinated afferent fibres innervating the primate skin and their 

response to noxious stimuli. J. Physiol. (Lond.) 197: 593–615, 1968. 

PETERSEN, M.AND LAMOTTE, R. H. Relationships between capsaicin sensitivity 

of mammalian sensory neurons, cell size and type of voltage gated 

Ca-currents. Brain Res. 561: 20–26, 1991. 

PETERSEN, M., LAMOTTE, R. H., KLUSCH, A., AND KNIFFKI, K. D. Multiple 

capsaicin-evoked currents in isolated rat sensory neurons. Neuroscience 75: 

495–505, 1996. 

REICHLING, D.B.AND LEVINE, J. D. Heat transduction in rat sensory neurons 

by calcium-dependent activation of a cation channel. Proc. Natl. Acad. Sci. 

USA 94: 7006–7011, 1997. 

RITTER, A. M., LEWIN, G.R.,AND MENDELL, L. M. Regulation of myelinated 

nociceptor function by nerve growth factor in neonatal and adult rats. Brain 

Res. Bull. 30: 245–249, 1993. 

SOMERO, G. N. Proteins and temperature. Annu. Rev. Physiol. 57: 43–68, 1995. 

SZALLASI, A. The vanilloid (capsaicin) receptor: receptor types and species 

differences. Gen. Pharmacol. 25: 223–243, 1994. 

SZALLASI, A.AND BLUMBERG, P. M. Vanilloid receptors: new insights enhance 

potential as a therapeutic target. Pain 68: 195–208, 1996. 

SZOLCSÁNYI, J., ANTON, F., REEH, P.W.,AND HANDWERKER, H. O. Selective 

excitation by capsaicin of mechano-heat sensitive nociceptors in rat skin. 

Brain Res. 446: 262–268, 1988. 

TILLMAN, D.-B., TREEDE, R.-D., MEYER, R.A.,AND CAMPBELL, J. N. Response 

of C fibre nociceptors in the anaesthetized monkey to heat stimuli: estimates 

of receptor depth and threshold. J. Physiol. (Lond.) 485: 753–765, 1995. 

TOMINAGA, M., CATERINA, M. J., MALMBERG, A. B., ROSEN, T. A., GILBERT, H., 

SKINNER, K., RAUMANN, B. E., BASBAUM, A.I.,AND JULIUS, D. The cloned 

capsaicin receptor integrates multiple pain-producing stimuli. Neuron 21: 

531–543, 1998. 

TREEDE, R.-D., MEYER, R.A.,AND CAMPBELL, J. N. Myelinated mechanically 

insensitive afferents from monkey hairy skin: heat response properties. 

J. Neurophysiol. 80: 1082–1093, 1998. 

TREEDE, R.-D., MEYER, R. A., RAJA, S.N.,AND CAMPBELL, J. N. Evidence for 

two different heat transduction mechanisms in nociceptive primary afferents 

innervating monkey skin. J. Physiol. (Lond.) 483: 747–758, 1995. 

VYKLICKY, L.AND KNOTKOVÁ-URBANCOVÁ, H. Can sensory neurones in culture 

serve as a model of nociception? Physiol. Res. 45: 1–9, 1996. 

WADDELL, P.J. AND LAWSON, S. N. Electrophysiological properties of subpopulations 

of rat dorsal root ganglion neurons in vitro. Neuroscience 36: 

811–822, 1990. 



on March 31, 2010

Treede Timo Kirschstein, Wolfgang Greffrath, Dietrich Büsselberg ...

Create successful ePaper yourself

Delete template?

Save as template?