This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a colleague Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Sommers, M. G. Articles by van Egmond, J. Search for Related Content PubMed PubMed Citation Articles by Sommers, M. G. Articles by van Egmond, J. Related Collections Mechanisms Pain Mechanisms Pain

---

ANALGESIA

Suppression of Noxious-Induced C-Fos Expression in the Rat Lumbar Spinal Cord by Isoflurane Alone or Combined with Fentanyl

Mathieu G. Sommers, DVM^*, Nha-Khanh Nguyen, MSc, Jan G. Veening, PhD, Kris C. Vissers, MD, PhD, FIPP, Merel Ritskes-Hoitinga, DVM, PhD, Dipl ECLAM^*, and Jan van Egmond, PhD

From the *Central Animal Laboratory, Department of Anatomy, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands; Department of Psychopharmacology, UIPS, Utrecht University, Utrecht, The Netherlands; and Department of Anesthesiology, Pain and Palliative Medicine, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.

Address correspondence and reprint requests to Mathieu G. Sommers, DVM, P.O. Box 9101, 231 CDL, NL-6500 HB Nijmegen, The Netherlands. Address e-mail to M.Sommers{at}cdl.umcn.nl.

Abstract

BACKGROUND: Although our understanding of nociceptive processing during anesthesia has increased greatly over the last decade, many patients still experience hyperalgesia and acute pain postoperatively. The noxious-induced withdrawal reflex (NIWR) model is specifically designed and validated to quantitatively study the reaction on painful, multimodal stimuli in animals under anesthetic conditions. Since the anesthetic mechanisms differ between inhaled anesthetics and opioids, we evaluated the differential effects of isoflurane and fentanyl on c-fos expression at the lumbar level as a measure of nociceptive information transfer during general anesthesia.

METHODS: The experimental setup consisted of a randomized block design with four experimental groups: two light minimum alveolar concentration (MAC) isoflurane anesthesia groups (unstimulated/NIWR-stimulated) and two NIWR-stimulated surgical anesthesia groups (1 MAC isoflurane anesthesia and MAC isoflurane anesthesia combined with fentanyl 400–600 µg · kg^–1 · h^–1). After 2 h of intermittent electrical stimulation of the hind paw of the rat, the number of Fos immunoreactive (Fos-IR) neurons in the dorsal horn was measured quantitatively.

RESULTS: The main suppressive effects on lumbar c-fos expression of isoflurane were observed in the superficial lamina II (P = 0.02), whereas fentanyl showed the strongest effects in lamina V (P = 0.05).

CONCLUSIONS: This study demonstrates that the NIWR model combined with spinal Fos-immunoreactivity is a suitable and useful model for evaluating the differential effects of inhaled anesthetics and opioids on nociceptive information transfer during general anesthesia.

Although our understanding of nociceptive processing, pain regulatory mechanisms, and effective pain management has increased greatly over the last decade,1 many patients still experience hyperalgesia and suboptimal pain control postoperatively.2 Further investigations, therefore, are essential to increase our understanding of the neuroplasticity evoked by noxious processing during anesthesia and to use this knowledge for improving intraoperative anesthetic, and especially analgesic depth assessment, thereby ameliorating negative postoperative outcome for patients undergoing surgery.3,4

Acute surgical pain models are useful to study nociception and the development of central sensitization at the spinal level. Many different models are available and each has its specific advantages and disadvantages.5 Ideal model characteristics for mimicking spinal neuroplastic changes caused by surgical stimulation would include intense, repetitive, multimodal noxious stimulation over prolonged periods. The noxious-induced withdrawal reflex (NIWR) model is an animal model specifically designed and validated to quantitatively study the reaction on painful, multimodal stimuli in animals under anesthetic conditions and therefore provides an indirect index of analgesia.6,7 Additionally, the NIWR correlates closely with depth of anesthesia.8 The electrical NIWR stimulation mimics the multimodal excitation pattern of different fiber types9 evoked by surgical incision and subsequent wound manipulations.

In pain research, Fos immunoreactivity (Fos-IR) is often used as a quantitative neuronal marker for spinal nociceptive information processing.10,11 The proto-oncogene, c-fos, is rapidly activated after noxious stimulation to express the protein, Fos, in neurons situated in spinal laminae and involved in nociceptive information transfer. Mapping c-fos expression in these neurons is at present the best global marker for efficiently locating individual neurons that respond to nociceptive input.

To validate the NIWR model for the study of central spinal sensitization during general anesthesia, we chose isoflurane and fentanyl as representatives of two frequently used drug groups in general anesthesia, namely inhaled anesthetics and opioids.

Since the anesthetic mechanisms differ between inhaled anesthetics and opioids12,13 and since their combination may result in a synergistic anesthetic action, the aim of the present study was to evaluate the differential effects of isoflurane alone or combined with fentanyl on c-fos expression at the lumbar level as a measure of nociceptive information transfer during general anesthesia. The absence of the hind paw withdrawal reflex was used as a reference level of anesthesia. Two different anesthesic protocols were investigated that fully suppressed the withdrawal reflex, namely a high-dose of isoflurane protocol and a low-dose of isoflurane plus fentanyl protocol.

METHODS

This study was approved by the Institutional Animal Care and Use Committee of the Radboud University Nijmegen and was conducted in accordance with the guidelines of the EC Council Directive 86/609/EEC. Twenty male Wistar rats were obtained from the SPF breeding facility of the Radboud University of Nijmegen. They weighed an average of 325 g and were group housed under controlled conditions (12 h light/dark cycle, lights on at 7:00 am, 21°C ± 2°C) and allowed to acclimatize for 2 wk. They received water ad libitum and standard rat chow (Ssniff R/MH, Ssniff Spezialdiäten Gmbh, Soest, Germany).

Experimental Design
Experimental setup consisted of a randomized block design. Four experimental groups were defined: The LIN group (n = 4) was not stimulated under light MAC (1.1%) isoflurane anesthesia: Low-dose Isoflurane Not stimulated. The LIS group (n = 6) was NIWR-stimulated under light MAC isoflurane anesthesia: Low-dose Isoflurane Stimulated. The HIS group (n = 4) was NIWR-stimulated under surgical 1 MAC (2.2%) isoflurane anesthesia: High-dose Isoflurane Stimulated. The LIFS group (n = 6) was NIWR-stimulated under surgical MAC isoflurane anesthesia combined with a continuous rated infusion (CRI) of fentanyl: Low-dose Isoflurane plus Fentanyl Stimulated. The fentanyl infusion was started 30 min before the NIWR stimulation series and was individually titrated to fully suppress the NIWR, leading to infusion rates ranging from 400 to 600 µg · kg^–1 · hr^–1. The choice for the low and high isoflurane anesthesia regimens was based on the MAC in rats (MAC₅₀ = 1.45%), leading to a MAC (1.1%) light baseline anesthesia and a 1 MAC (2.2%) surgical anesthesia.

In all groups, anesthesia was induced with 5% isoflurane in an oxygen/air mixture (Fio₂ = 0.4) and continued at 2% isoflurane during the preparation period. All rats were placed in the stimulation setup in lateral position after tracheal intubation, IV access into the tail vein and application of the electrical stimulation shoe on their left hind paw. Isoflurane was adjusted to 1.1% for the LIS, LIFS, and LIN groups and to 2.2% for the HIS group. Body temperature (37.0°C ± 0.5°C, dedicated equipment), ventilation (4.0% ± 0.5% end tidal CO₂, QP9000 mass spectrometer, CaSE Ltd., Biggin Hill, UK), and oxygen saturation (95%, Nonin 8600V, Nonin Medical Inc, Plymouth, MN) were controlled. Animals in all groups received IV fluid support by a CRI at 8 mL^–1 · kg^–1 · h^–1, to which fentanyl was added only for animals in the LIFS group. After 30 min acclimatization in the experimental setup, NIWR stimulation was started.

The NIWR Model
An electrical multimodal noxious stimulation was applied by the stimulation shoe on the hind paw of the rat and the isometrically developed tension of the withdrawal reflex was measured quantitatively with a calibrated force displacement transducer (TB-611T, Nihon Kohden, Tokyo, Japan) as a clinical measure for depth of anesthesia (in-house developed data acquisition system). The NIWR stimulation protocol consisted of a 500 ms train of electrical block pulses (100 Hz, 7.5 mA, 4 ms) that was repeated every 80 s for 2 h (Grass CCU1 Constant Current and SIU5 Isolation unit, Grass Stimulator S88, Grass Technologies, the Netherlands). Adequate depth of anesthesia for the HIS and LIFS groups was defined as a continuing absence of the withdrawal reflex after NIWR stimulation.

Fos Immunostaining
Subsequent to the 2 h stimulation period, anesthesia was quickly deepened with 5% isoflurane and the animals were perfused via the heart with 75–100 mL saline, followed by 400 mL of cold paraformaldehyde in phosphate buffered saline (4% paraformaldehyde in 0.1 M PBS, pH 7.3), followed by removal of the spinal cord. After overnight postfixation in the same fixative at 4°C, spinal cords were stored in cold buffered (0.1 M PBS) 30% sucrose. Frozen cross-sections of the lumbosacral part of the spinal cord (40 µm thickness) were collected in 6 parallel series, and stored in cold 0.1 M PBS.

At least one series of free-floating sections per animal was used for immunocytochemical staining for Fos-IR. (Antibody: polyclonal anti-Fos antiserum, raised in rabbit, (Santa Cruz Biotechnology Inc., Santa Cruz, CA) diluted 1:20,000, room temperature, overnight). After the second incubation of 90 min in donkey anti-rabbit antiserum conjugated to peroxidase (1:1500, Jackson Immunoresearch, Westgrove, PA), sections were incubated for 90 min in Vector ABC-elite (1:800) (Vectastain, Brunschwig Chemie, Amsterdam, the Netherlands). The Fos-antibody peroxidase complex was visualized by a 0.02% 3,3'-diaminobenzidine tetrahydrochloride (DAB; Sigma Aldrich, St. Louis, MO), followed by a 0.3% Ni-ammonium sulfate and 0.006% hydrogen peroxide treatment for a blue-black staining of Fos-IR. Sections were extensively rinsed with 0.1 M PBS between the successive steps.

All sections of a series between spinal levels L2 and S2 were mounted on gelatin-coated object glasses and dried overnight. After dehydration in an alcohol series, sections were cleared in Xylol and mounted in Entellan. Further details of the immunocytochemical staining procedures have been described previously.14,15

Quantification Procedures
Quantification of the caudal L4 lumbar sections occurred by application of a grid square (40 µm for lamina I, 80 µm for laminae II, V, and X). Counting of Fos-IR occurred in three grid squares at a magnification of 10 x 20. For each rat, the caudal lumbar L4 section with the most Fos-IR was used for this quantification procedure. The spinal gray matter was divided into Rexed's laminae, according to Molander et al.16 and the rat brain atlas of Paxinos & Watson,17 as illustrated for the lumbar dorsal horn at the L4 level in Figure 1. In preceding pilot-experiments, it was observed that the main changes in Fos-IR could be observed in laminae I, II, V, and X. These laminae were selected for quantification and inside the selected laminae the location of the selected squares were in the medial part of lamina I, and in the medial parts of laminae II and V. From Figure 3, it is immediately clear that most of the Fos-IR was detected in these regions of the laminae. This quantification method has been described extensively.14 The Fos-IR count can be used to measure relative changes in neuronal activity per spinal lamina, but does not represent absolute numbers of activated neurons. After quantifying the numbers of Fos-immunoreactive neurons, the statistical analysis in SPSS 12.0.1 of the numbers obtained in the different groups comprised an Univariate Kruskal–Wallis ANOVA (analysis of variance) test, followed by Mann–Whitney U-test post hoc analysis, with 5% significance levels. Group results are represented as mean (sem) Fos-IR neurons per three grid-squares.

View larger version (26K): [in this window] [in a new window]   Figure 1. Reference figure to illustrate the laminar organization of the dorsal horn of the rat spinal cord at level L4.

View larger version (126K): [in this window] [in a new window]   Figure 3. Microphotographs of Fos-IR at the caudal L4 level for all experimental groups. A, the amount of Fos-IR in the ipsilateral dorsal horn in the low-dose isoflurane stimulated (LIS) group: a high density in the superficial layers and numerous activated neurons in the deeper layers; the insert shows a detail of the superficial activation, similar to inserts in B, C, and D; B, the activation in the high-dose isoflurane stimulated (HIS) group; activation in the superficial layers is decreased but in the deeper layers, activation is still considerable; C, the neuronal activation in the low-dose isoflurane plus fentanyl stimulated (LIFS) group; the density of Fos-IR in the superficial layers has hardly changed, but in the deeper layers less activation is present; D, the activation in the low-dose isoflurane not stimulated (LIN) group; only a few Fos-IR neurons can be detected. See legend of Figure 2 for further group details.

View larger version (46K): [in this window] [in a new window]   Figure 2. Representative examples of 20 min noxious-induced withdrawal reflex (NIWR) recordings (a total of 15 NIWR stimulations every 80 s) for all experimental groups. LIS = low-dose isoflurane stimulated; LIFS = low-dose isoflurane plus fentanyl stimulated; HIS = high-dose isoflurane stimulated; LIN = low-dose isoflurane not stimulated. Noxious induction of the withdrawal reflex as measured by the isometrically developed tension is illustrated for LIS, suppression of NIWR is illustrated for LIFS and HIS, stability of NIWR baseline is illustrated for LIN.

Of the total study group of 20 animals, one animal of the HIS group was excluded because tissue quality was inadequate for Fos-IR staining.

The NIWR stimulation induced a robust withdrawal reflex in the LIS group and an absent withdrawal reflex in the LIN group (Fig. 2). Suppression of the NIWR in the HIS group was very effective. Full suppression of the NIWR in the LIFS group required high fentanyl infusion rates ranging from 400 to 600 µg · kg^–1 · h^–1.

In the LIS group (Fig. 3A), NIWR stimulation resulted in a reproducible pattern of Fos-IR ipsilaterally in the lumbosacral spinal cord (Fig. 4). The highest density of Fos-IR neurons was present in the medial part of the superficial laminae I and II of the L4–5 lumbar segments. Also, in lamina V of the dorsal horn, a substantial activation was observed and, to a lesser extent, in the intermediate dorsal horn layers and in lamina X.

View larger version (15K): [in this window] [in a new window]   Figure 4. Differential Fos-IR expression for all experimental groups, clustered per spinal lamina. In the high-dose isoflurane stimulated (HIS) group, the number of noxious-induced Fos-IR neurons was reduced overall in the dorsal horn laminae examined and specifically in lamina II compared with the low-dose isoflurane stimulated (LIS) group. The HIS group of Fos-IR neurons also was reduced in lamina II compared with the low-dose isoflurane plus fentanyl stimulated (LIFS) group. The LIFS group was not different overall, but was reduced in lamina V compared with the LIS group. See legend Figure 2 for further group details. Significance a: P 0.05 compared with low-dose isoflurane not stimulated (LIN); b: P 0.05 compared with LIS; c: P 0.05 for LIFS versus HIS.

In the HIS group (Fig. 3B), the overall number of noxious-induced Fos-IR neurons was significantly reduced in the examined dorsal horn laminae (Fig. 4) compared with the LIS group (38 vs 12 Fos-IR neurons, P = 0.02). In lamina II, the Fos-IR was significantly decreased compared with the LIS group (5 vs 17 Fos-IR neurons, P = 0.02) and compared with the LIFS group (5 vs 12 Fos-IR neurons, P = 0.02).

In the LIFS group (Fig. 3C), the overall number of noxious-induced Fos-IR neurons was not reduced (Fig. 4). However, in lamina V, the Fos-IR was significantly diminished only in the LIFS group compared with the LIS group (5 vs 11 Fos-IR neurons, P = 0.05).

In the LIN group (Fig. 3D), a few Fos-IR neurons could be detected in the superficial laminae of the dorsal horn (Fig. 4).

In all groups, in the ipsilateral ventral horn and all contralateral laminae, only a few Fos-IR neurons were present in all groups (data not shown).

In conclusion, the main effects of isoflurane were observed in the superficial laminae, whereas fentanyl showed the strongest effects in lamina V (Fig. 4).

DISCUSSION

This study demonstrates for the first time that the combination of the NIWR model and lumbar Fos expression is a suitable and useful model for studying nociceptive information transfer at the spinal level during anesthesia. The finding that the suppression pattern of Fos-IR in the dorsal horn laminae differs between the HIS and LIFS groups is new. A high-dose of isoflurane preferentially decreased Fos-IR in laminae I and II, whereas the combination of a low-dose of isoflurane and fentanyl preferentially diminished Fos-IR in lamina V.

The NIWR stimulus is electrical. Therefore, typical positive properties5 are that it is a quantifiable, reproducible, and noninvasive stimulus, and typical negative properties5 are that it excites peripheral fibers in a nonspecific fashion, including non-noxious fibers, and that it bypasses peripheral receptors. The nonspecific excitation ensures that all noxious fiber types are included, which can hardly be arranged with a natural stimulus. However, non-noxious large diameter fibers are more easily excited, and so supramaximal electrical noxious excitation is always accompanied by supramaximal non-noxious excitation, which may activate inhibitory spinal mechanisms before arrival of noxious action potentials in the spinal cord. Fortunately, electrical tetanic stimulation comparable with NIWR stimulation is experienced as painful in humans.18,19 It is also important to note that electrical stimulation will induce maximal spinal c-fos when stimulation intensity is at A/C-fiber strength and will induce minimal spinal c-fos when stimulation intensity is at A/Aβ strength.20–22 The bypassing of peripheral receptors is an advantage in our study, since it eliminates the bias of inhibitory (opioid) effects on peripheral nociceptors. This effect could otherwise diminish spinal Fos expression.

The assessment of adequate anesthetic and analgesic depth in this study is based on the absence of the hind paw withdrawal reflex. The spinal withdrawal reflex arch consists of dorsal horn projection neurons, interneurons, and ventral horn motor neurons. Therefore, one can assume that the assessment of anesthetic depth is better correlated with the withdrawal reflex than the assessment of analgesic depth, which involves the dorsal horn area in particular. This may partly explain why high fentanyl CRI was needed in addition to MAC isoflurane to achieve sufficient anesthetic depth, which may already have been achieved at lower fentanyl infusion rates. However, our anesthetic end-point in this study well illustrates that, in contrast with high isoflurane protocols, low isoflurane plus high opioid CRI protocols could leave superficial spinal laminae sensitive for peripheral input, which may contribute to postoperative hyperalgesia. In future studies, this hypothesis may be assessed using von Frey hyperalgesia testing after anesthetic recovery.

Our findings on the suppressive effects on Fos-IR of the combination of isoflurane and fentanyl are new, and can therefore be compared only with studies investigating the effects of each drug separately.

The HIS effects on lamina I/II are in accordance with the study of Jinks et al., wherein Fos-IR induced by mechanical clamping was measured during isoflurane or halothane anesthesia.23 These authors observed that overall lumbar dorsal horn Fos-IR was suppressed in the high isoflurane group, including superficial laminae I/II and lamina V. Their percentual suppression in laminae I/II and V was larger than in the present study, probably because our stimulation was more intense and of longer duration. The limited LIFS effects on lamina I/II and extensive suppressive effects on lamina V are in accordance with subplantar formalin injection studies24,25 and a noxious heat study.26 In the first formalin study, Fos-IR was more decreased in lamina V than in laminae I + II by pretreatment with a fentanyl 0.5 µg dosage intrathecally. In the second formalin study, a preferential Fos-IR decrease in lamina V was found using the formalin model with a fentanyl 100 µg/kg dosage.25

In summary, the NIWR model combined with spinal Fos-IR is a suitable model for evaluating the differential effects of inhaled anesthetics and opioids on nociceptive information transfer during general anesthesia. The suppression of Fos-IR by isoflurane and fentanyl in the spinal laminae of the lumbar dorsal horn are in accordance with other studies using different stimulus modalities.

Further investigations are needed to broaden the scope of the combination of the NIWR model and c-fos on nociceptive processing during anesthesia. First, by evaluating effects of other anesthetics and analgesics and, second, by extending the c-fos analysis to supraspinal nuclei involved in nociceptive pathways. This may contribute to a better understanding of the relation between anesthetic depth assessment and the suppression of nociceptive systems during surgery.

ACKNOWLEDGMENTS

We acknowledge Mrs. F. van de Pol for her biotechnical assistance and Mr. P.J. Dederen for his assistance with the immunohistochemical procedures.

Footnotes

Accepted for publication December 3, 2007.

REFERENCES

1. Edwards RR. Individual differences in endogenous pain modulation as a risk factor for chronic pain. Neurology 2005; 65:437–43[Abstract/Free Full Text]
2. Wilder-Smith OH, Arendt-Nielsen L. Postoperative hyperalgesia: its clinical importance and relevance. Anesthesiology 2006;104:601–7[Web of Science][Medline]
3. Brennan TJ, Kehlet H. Preventive analgesia to reduce wound hyperalgesia and persistent postsurgical pain: not an easy path. Anesthesiology 2005;103:681–3[Web of Science][Medline]
4. Woolf CJ, Salter MW. Neuronal plasticity: increasing the gain in pain. Science 2000;288:1765–9[Abstract/Free Full Text]
5. Le Bars D, Gozariu M, Cadden SW. Animal models of nociception. Pharmacol Rev 2001;53:597–652[Abstract/Free Full Text]
6. Duysens J, Gybels J. The automated measurement of hind limb flexor reflex of the rat as a substitute for the tail-flick assay. J Neurosci Methods 1988;24:73–9[Web of Science][Medline]
7. Duysens J, Dom R, Gybels J. Suppression of the hind limb flexor reflex by stimulation of the medial hypothalamus and thalamus in the rat. Brain Res 1989;499:131–40[Web of Science][Medline]
8. van den Broek PL, van Rijn CM, van Egmond J, Coenen AM, Booij LH. An effective correlation dimension and burst suppression ratio of the EEG in rat. Correlation with sevoflurane induced anaesthetic depth. Eur J Anaesthesiol 2006;23:391–402[Web of Science][Medline]
9. Chang C, Shyu BC. A fMRI study of brain activations during non-noxious and noxious electrical stimulation of the sciatic nerve of rats. Brain Res 2001;897:71–81[Web of Science][Medline]
10. Coggeshall RE. Fos, nociception and the dorsal horn. Prog Neurobiol 2005;77:299–352[Web of Science][Medline]
11. Harris JA. Using c-fos as a neural marker of pain. Brain Res Bull 1998;45:1–8[Web of Science][Medline]
12. Sonner JM, Antognini JF, Dutton RC, Flood P, Gray AT, Harris RA, Homanics GE, Kendig J, Orser B, Raines DE, Rampil IJ, Trudell J, Vissel B, Eger EI. Inhaled anesthetics and immobility: mechanisms, mysteries, and minimum alveolar anesthetic concentration. Anesth Analg 2003;97:718–40[Abstract/Free Full Text]
13. Inturrisi CE. Clinical pharmacology of opioids for pain. Clin J Pain 2002;18:S3–S13[Web of Science][Medline]
14. Coolen LM, Peters HJ, Veening JG. Anatomical interrelationships of the medial preoptic area and other brain regions activated following male sexual behavior: a combined fos and tract-tracing study. J Comp Neurol 1998;397:421–35[Web of Science][Medline]
15. Veening JG, Bouwknecht JA, Joosten HJ, Dederen PJ, Zethof TJ, Groenink L, van der Gugten J, Olivier B. Stress-induced hyperthermia in the mouse: c-fos expression, corticosterone and temperature changes. Prog Neuropsychopharmacol Biol Psychiatry 2004;28:699–707[Medline]
16. Molander C, Xu Q, Grant G. The cytoarchitectonic organization of the spinal cord in the rat. I. The lower thoracic and lumbosacral cord. J Comp Neurol 1984;230:133–41[Web of Science][Medline]
17. Paxinos G, Watson C. The rat brain in stereotaxic coordinates. 5th ed. Academic Press, 2005
18. Baars JH, Kalisch D, Herold KF, Hadzidiakos DA, Rehberg B. Concentration-dependent suppression of F-waves by sevoflurane does not predict immobility to painful stimuli in humans. Br J Anaesth 2005;95:789–97[Abstract/Free Full Text]
19. Rehberg B, Grunewald M, Baars J, Fuegener K, Urban BW, Kox WJ. Monitoring of immobility to noxious stimulation during sevoflurane anesthesia using the spinal H-reflex. Anesthesiology 2004;100:44–50[Web of Science][Medline]
20. Herdegen T, Kovary K, Leah J, Bravo R. Specific temporal and spatial distribution of JUN, FOS, and KROX-24 proteins in spinal neurons following noxious transsynaptic stimulation. J Comp Neurol 1991;313:178–91[Web of Science][Medline]
21. Hunt SP, Pini A, Evan G. Induction of c-fos-like protein in spinal cord neurons following sensory stimulation. Nature 1987;328:632–4[Medline]
22. Herdegen T, Leah JD. Inducible and constitutive transcription factors in the mammalian nervous system: control of gene expression by Jun, Fos and Krox, and CREB/ATF proteins. Brain Res Brain Res Rev 1998;28:370–490[Medline]
23. Jinks SL, Antognini JF, Martin JT, Jung S, Carstens E, Atherley R. Isoflurane, but not halothane, depresses c-fos expression in rat spinal cord at concentrations that suppress reflex movement after supramaximal noxious stimulation. Anesth Analg 2002; 95:1622–8[Abstract/Free Full Text]
24. Nakamura T, Takasaki M. Intrathecal pre-administration of fentanyl effectively suppresses formalin evoked c-Fos expression in spinal cord of rat. Can J Anaesth 2001;48:993–9[Web of Science][Medline]
25. Lin FS, Shyu BC, Shieh JY, Sun WZ. Nitrous oxide suppresses tonic and phasic nociceptive behaviors but not formalin-induced c-Fos expression in the rat spinal cord dorsal horn. Acta Anaesthesiol Sin 2003;41:115–23[Medline]
26. Tolle TR, Herdegen T, Schadrack J, Bravo R, Zimmermann M, Zieglgansberger W. Application of morphine prior to noxious stimulation differentially modulates expression of Fos, Jun and Krox-24 proteins in rat spinal cord neurons. Neuroscience 1994;58:305–21[Web of Science][Medline]

This article has been cited by other articles:

				J. L. P. Giele, A. F. Nabers, J. G. Veening, J. van Egmond, and K. C. P. Vissers The Effect of a Thoracic Spinal Block on Fos Expression in the Lumbar Spinal Cord of the Rat Induced by a Noxious Electrical Stimulus at the Hindpaw Anesth. Analg., November 1, 2009; 109(5): 1659 - 1665. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a colleague Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Sommers, M. G. Articles by van Egmond, J. Search for Related Content PubMed PubMed Citation Articles by Sommers, M. G. Articles by van Egmond, J. Related Collections Mechanisms Pain Mechanisms Pain