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Division of Pain Medicine, Department of Anesthesiology, Perioperative Medicine and Pain Management, Jackson Memorial Hospital/University of Miami, Miami, Florida
Address correspondence and reprint requests to Salahadin Abdi, MD, PhD, Director, MGH Pain Center, 15 Parkman Street, WACC 333B, Boston, MA 02114. Address e-mail to sabdi{at}partners.org
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Abstract |
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Introduction |
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The anatomy of the SG in rats closely resembles that of humans. In humans, the cervical sympathetic chain is composed of superior, middle, and inferior cervical ganglia. The inferior cervical ganglion is often fused with the first thoracic sympathetic ganglion, forming the SG. It is generally located anterior to the first rib and extends between C7 and T1 interspace. In rats, the SG consists of the inferior cervical ganglion and the first 3 thoracic sympathetic ganglia (C6-T3) fused together and is located at the level of the first 2 thoracic vertebrae (2,3). Consequently, it is possible to simulate SGB in an experimental rat model.
Although SGB is routinely used in the clinical setting for chronic pain patients, it is unclear if this block affects myocardial contractility and hemodynamics. The few published studies in rats and humans report contradictory findings. All the previous studies in rats accessed the ganglion through a thoracotomy approach, which is invasive and painful. The purpose of the present study was to establish a simple, less painful and reproducible technique of SGB in rats.
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Methods |
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Twenty-six female Sprague-Dawley rats were divided into 6 groups as follows: Group I (n = 4) had right sided SGB; Group II (n = 5) had left SGB; Group III (n = 5) had bilateral SGB. Three corresponding sham groups (Groups IV–VI; n = 4 in each group) were used as controls.
The rats were endotracheally intubated after induction of general anesthesia with isoflurane (2.5 vol% for induction and 1 vol% for maintenance) in oxygen and mechanically ventilated with a rodent ventilator (Rodent ventilator 683; Harvard Apparatus, Cambridge, MA). The body temperature was kept at a constant 37°C using a heating pad.
Electrodes for monitoring heart rate and rhythm were connected to the animals and the electrocardiogram (ECG) data were collected using an ECG amplifier (CP 511 A.C. amplifier, Astro-med, Grass Group) and computer software (Grass Polyview Data Acquisition and Analysis System) for real-time or retrograde analysis. Baseline measurements were done after a stabilization period of 60 min, at which time local anesthetics or saline injection into the SG was done via our new posterior percutaneous technique. Treatment groups received 0.2 mL of 0.25% bupivacaine HCL whereas control (sham groups) received 0.2 mL saline.
The cartilaginous process of C7 spinous process (CPSP) was used as a landmark. This is located between the two scapulae and is elevated approximately 0.5 cm from the surrounding tissue (Figure 1). This structure was palpated and a short beveled needle (25-gauge 5/8" needle) attached with a 1-mL syringe was inserted in a paramedian sagittal approach and advanced in a posterioanterior direction along the side of the C7 vertebral body. When the tip of the needle lost contact with the vertebral body (a sign that it has passed the anterior aspect of the vertebral body) it was withdrawn approximately 0.5 mm and the local anesthetics (0.2 mL of 0.25% bupivacaine HCL for each unilateral block) were injected. The rats were under light general anesthesia during the procedure. The rats recovered from the general anesthesia within a few minutes after the block and thus could be observed for the development of ptosis.
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The degree and extent of ptosis was evaluated and recorded by a person who was blinded as to the study as follows:
– no difference of eye size between the two sides (no ptosis).
± unsure about the difference in eye size between the two sides (unclear) ptosis.
+ barely noticeable upper eyelid droop (mild ptosis).
++ eye size in blocked side was more than half of the contralateral one (moderate ptosis).
+++ eye size in blocked side was less than half of the contralateral one (severe ptosis).
Heart rate data were collected at baseline (before), 5, 10, 15, 20, 30, 45, 60, and 90 min after SGB. The values at different time points were compared within and between groups at different time points.
Nine female Sprague-Dawley rats were used to verify the accuracy of our new technique. The SGs of the animals (right SG; n = 3, left SG; n = 3 and bilateral SG; n = 3) were injected with 0.2 mL of 0.5% methylene blue as described above. Immediately thereafter, the animals were killed and their chests were opened under the microscope (PZMTIII, World Precision Instruments, Sarasota, FL). The ipsilateral and contralateral SGs were assessed for methylene blue staining.
All data are expressed as mean ± sem. GraphPad Prism statistic analysis software was used for the statistical evaluation (GraphPad, San Diego, CA). One-way analysis of variance was used to compare baseline heart rates at different time points within the same group and Student’s t-test was used to compare the values among groups. Statistical significance was defined at P < 0.05.
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Results |
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There was no statistically significant change in heart rate after unilateral SGB neither among groups nor within the groups. However, bilateral SGB resulted in a significant reduction in heart rate, which was observed as early as 5 min post-block and lasted for 45 min (Fig. 3).
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Twelve SGs of 9 rats were injected with 0.5% methylene blue using our new technique. At the autopsy, all of the SGs except one (11 of 12) were stained with methylene blue (Fig. 4). In one case, the methylene blue stained the pectoral muscle in the axillary area, probably as a result of an oblique or medio-lateral direction of the inserted needle. Unilateral methylene blue injection did not result in a contralateral stain.
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Discussion |
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Ptosis is generally accepted as one of the main indicators of a clinically successful SGB. We thus recorded the timecourse and the degree of its development after the block. Furthermore, we injected methylene blue using the same percutaneous technique to verify the reliability of this newly developed technique. We were successful in staining 11 of the 12 ganglions with methylene blue. Our results show that our new technique of SGB is a reliable and simple method to block the SG in rats.
In the clinical setting the development of Horner’s syndrome after the performance of SGB is a valuable sign of a successful sympathetic blockade (5). Dellemijn et al. (6) reported that all their study patients who received SGB developed Horner’s syndrome and nasal stuffiness, which was associated with pain relief. The authors further reported that changes in skin temperature after the SGB did not correlate with pain relief. Another study which reported a discrepancy between changes in skin temperature and SGB was published by Stevens et al. in 1998 (7). The authors concluded from their study that SGB often failed to increase skin temperature in the ipsilateral hand. These clinical observations are partly similar to our laboratory findings using rats; namely, we found that ptosis was observed in all the animals that underwent SGB. This phenomenon lasted for up to 30 minutes. We also found that it was difficult to monitor miosis and enophthalmos in rats, which is in contrast to humans. Thus our findings are consistent with those of Hanamatsu et al. (4).
The effect of SGB on changes in heart rate in humans remains unstudied. In fact, the literature concerning changes in ECG and heart rates induced by unilateral SGB on normal subjects is controversial. Rogers et al. (8) noted lateralization of sympathetic control of the human sinus node, which results in a decrease in heart rate after right-sided SGB but not after left SGB. In addition, Kashima et al. (9) demonstrated a significant increase in the P-P interval post-right SGB in normal subjects. Conversely, Fujiki et al. (10) reported that neither right nor left SGB induced any significant change in both RR and corrected QT intervals. Pardini et al. (11) injected a fluorescent retrograde tracer into either the left or right ventricular free walls in rats and showed that approximately 92% of all labeled neurons were located bilaterally in the middle and inferior cervical ganglia. They concluded that bilateral middle and inferior cervical ganglionectomy would be expected to eliminate 92% of cardiac sympathetic postganglionic cell bodies. Their study also showed that up to 100% of norepinephrine stores were depleted by bilateral removal of the middle cervical-SG complex. In our rat model, we observed that heart rate significantly decreased for up to 45 minutes after bilateral SGB. However, unilateral SGB did not affect heart rate. This observation could be explained by a temporary depletion of norepinephrine stores, leading to a successful block of cardiac sympathetic activity.
There are several limitations to the present study. Although our new technique is simple and easy to learn, it has yet to be reproduced and evaluated by others. Furthermore, the physiologic variables (i.e., ptosis and temperature changes in the affected limb) associated with a successful SGB in rats are unknown. In the present study, we did not evaluate limb temperature changes; therefore, an incomplete block may be produced by this technique. Although clinically a fluoroscopically guided and precisely performed SGB may result in Horner’s syndrome without changes in ipsilateral skin temperature, this has not been verified in rats. Thus, a head-to-head comparison between the two variables (temperature increase versus Horner’s syndrome), possibly including ipsilateral regional blood flow changes and electrophysiological studies to verify the completeness of the block, might be beneficial to define a successful SGB in rats. Finally, this study was done on healthy rats and thus the efficacy of SGB in rats with forepaw nerve injury (neuropathic pain model) must be investigated.
In summary, we developed a simple and reliable technique of SGB in rats in which we used CPSP of C7 as a landmark. Finally, bilateral SGB can result in a significant bradycardia, which might or might not be beneficial to the patient. Our present rat model may be used for future studies about the role of cervical sympathetic nervous system (especially the SG) in regulating sympathetically maintained pain and myocardial function in rats.
We gratefully acknowledge Dr. Edir B. Siqueira and Mr. Bernard Jay Wasserlauf for their excellent assistance and advice in this project.
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Footnotes |
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Accepted for publication January 28, 2005.
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