Buprenorphine Disrupts Sleep and Decreases Adenosine Levels in Sleep-Regulating Brain Regions of Sprague Dawley Rat

doi:10.1097/ALN.0b013e31822e9f85

Anesthesiology. Author manuscript; available in PMC 2012 October 1.

Published in final edited form as:

Anesthesiology. 2011 October; 115(4): 743–753.

doi: 10.1097/ALN.0b013e31822e9f85

PMCID: PMC3197808

NIHMSID: NIHMS319694

Buprenorphine Disrupts Sleep and Decreases Adenosine Levels in Sleep-Regulating Brain Regions of Sprague Dawley Rat

Elizabeth A. Gauthier, B.S., Medical Student, Sarah E. Guzick, B.S., Graduate Student, Chad M. Brummett, M.D., Assistant Professor, Helen A. Baghdoyan, Ph.D., Professor, and Ralph Lydic, Ph.D., Professor

Department of Anesthesiology, University of Michigan Ann Arbor, MI

Corresponding author: Ralph Lydic, 7433 Medical Science I, 1150 West Medical Center Drive, Ann Arbor, MI 48109, Phone: 734-647-7831; Fax: 734-764-9332; Email: rlydic/at/umich.edu

The publisher's final edited version of this article is available free at Anesthesiology

Abstract

Background

Buprenorphine, a partial μ opioid receptor agonist and κ opioid receptor antagonist, is an effective analgesic. The effects of buprenorphine on sleep have not been well characterized. This study tested the hypothesis that an antinociceptive dose of buprenorphine decreases sleep and decreases adenosine levels in regions of the basal forebrain and pontine brain stem that regulate sleep.

Methods

Male Sprague Dawley rats were implanted with intravenous catheters and electrodes for recording states of wakefulness and sleep. Buprenorphine (1 mg/kg) was administered systemically via an indwelling catheter and sleep/wake states were recorded for 24 h. In additional rats buprenorphine was delivered by microdialysis to the pontine reticular formation and substantia innominata of the basal forebrain while simultaneously measuring adenosine.

Results

An antinociceptive dose of buprenorphine caused a significant increase in wakefulness (25.2%) and a decrease in both nonrapid eye movement sleep (−22.1%) and rapid eye movement sleep (−3.1%). Buprenorphine also increased electroencephalographic delta power during nonrapid eye movement sleep. Coadministration of the sedative/hypnotic eszopiclone diminished the buprenorphine-induced decrease in sleep. Dialysis delivery of buprenorphine significantly decreased adenosine levels in the pontine reticular formation (−14.6%) and substantia innominata (−36.7%). Intravenous administration of buprenorphine significantly decreased (−20%) adenosine in the substantia innominata.

Conclusions

Buprenorphine significantly increased time spent awake, decreased nonrapid eye movement sleep, and increased latency to sleep onset. These disruptions in sleep architecture were mitigated by coadministration of the nonbenzodiazepine sedative/hypnotic eszopiclone. The buprenorphine-induced decrease in adenosine levels in basal forebrain and pontine reticular formation is consistent with the interpretation that decreasing adenosine in sleep-regulating brain regions is one mechanism by which opioids disrupt sleep.

Introduction

Opioids are used effectively in the treatment of chronic and acute pain, and the extensive use of opioids encourages efforts to develop counter-measures to combat unwanted side effects.^1,2 Opioids disrupt sleep^3-7 and sleep disruption can contribute to hyperalgesia,^8-16 impaired immune function,¹⁷ and postoperative cognitive impairment.^18,19

Adenosine is an endogenous neuromodulator that significantly enhances sleep²⁰ and diminishes nociception.²¹ Sleep is increased by increasing adenosine in the pontine reticular formation (PnO)^22-24 and in the substantia innominata (SI) area of the basal forebrain.^20,25 Adenosine levels in the PnO and SI are decreased by the μ opioid receptor agonists morphine and fentanyl.²⁶

Buprenorphine, a partial μ opioid receptor agonist and κ opioid receptor antagonist, is an effective analgesic but no prior studies have quantified the effects of buprenorphine on sleep architecture^7,27,28 or on adenosine levels in the PnO and SI. This study was designed to test the hypothesis that buprenorphine decreases sleep and adenosine levels in PnO and SI, brain regions known to modulate sleep and nociception.

Materials and Methods

Animals

Adult, male Crl:CD^{^*}(SD) (Sprague Dawley) rats (n = 26) purchased from Charles River Laboratories (Wilmington, MA) were used for all studies. Rats weighing 250 to 350 g were used because brains from rats in this weight range are known to fit the rat stereotaxic atlas.²⁹ Male rats were chosen to facilitate comparison of the present results to previous data obtained from males.^26,30-32 Rats were housed in a 12:12-h light/dark cycle (lights on from 8:00 to 20:00) with access to food and water ad libitum. Procedures were reviewed and approved by the University of Michigan Committee on the Use and Care of Animals. Every phase of this study adhered to the Guide for the Care and Use of Laboratory Animals: Eighth Edition, National Academy of Sciences Press, Washington DC, 2011.^{^*}

Surgical Procedures

Rats were anesthetized with 3% isoflurane (Hospira, Inc., Lake Forest, IL). The jugular vein was exposed and a catheter (12 cm of Micro-Renathane tubing (MRE–040), Braintree Scientific, Braintree, MA) was inserted in the direction of the heart. The other end of the catheter was tunneled subcutaneously and implanted between the scapulas. A back-mounted flange guide cannula (8I 1000BM10, Plastics One, Roanoke, VA) and dummy cannula (8IC313DCCACC, Plastics One) were secured with the catheter in the midscapular position. This procedure provided subsequent venous access.

Implantation of the jugular vein catheter was immediately followed by implantation of electrodes for recording sleep. Rats were moved to a Kopf Model 962 small animal stereotaxic instrument fitted with a Model 906 rat anesthesia mask (David Kopf Instruments, Tujunga, CA) and anesthesia was maintained with 2.0 % isoflurane. Three electrodes (8IE36320SPCE, Plastics One) for recording cortical electroencephalogram were placed 2.0 mm posterior and 1.3 mm lateral to bregma, 2.0 mm posterior and 1.5 mm lateral to bregma, and 1.0 mm anterior and 1.5 lateral to bregma.²⁹ Two electrodes (4 cm of AG 7/40T Medwire, Mt. Vernon, NY) for electromyogram recordings were placed in the dorsal neck muscle, and a third electrode was placed under the skin of the neck muscle as a reference. The nonimplanted ends of the electroencephalogram and electromyogram electrodes were soldered to electrical contact pins (E363/0, Plastics One) that were plugged into a plastic pedestal (8K00022980IF, Plastics One). Three stainless steel anchor screws (MPX-0080-02P-C, Small Parts Inc., Miami Lakes, FL) were placed in the skull to secure the electrodes. Dental acrylic was used to construct a head cap covering the electrodes and to anchor the electrical connector and electrodes to the skull. Rats were then removed from the stereotaxic frame and monitored during recovery from anesthesia. Once ambulatory, animals were returned to their home cages.

Behavioral Conditioning for Sleep Recordings

Following 1 week for surgical recovery, rats were conditioned for an additional week to 10 days to sleeping in a Raturn (Bioanalytical Systems, West Lafayette, IN) recording chamber. During conditioning the implanted electrodes were attached by a cable (363–441/six 80CM 6TCM, Plastics One) to amplifiers and a computer for digital recording of electroencephalogram and electromyogram signals. Rats had free access to food and water while in the recording chambers.

Nociceptive testing

An initial series of experiments was conducted to confirm that the 1 mg/kg dose of buprenorphine produced anti-nociception as reported previously.³³ Procedures for thermal nociceptive testing have been described in detail.^34,35 Briefly, rats were conditioned to being placed in the Plexiglas chamber of a Hargreaves Paw Withdrawal unit (Model 336T, IITC Life Science, Woodland Hills, CA) one hour each day for the week prior to data collection. The Model 400 (IITC Life Science) heated glass stand and base was set to 30°C for the last ten minutes of each conditioning session and both hind paws of the rat were exposed to the heat stimulus.³⁶ Five baseline measurements were taken after the habituation time. As soon as baseline measurements were recorded, saline or buprenorphine hydrochloride (Sigma-Aldrich, Saint Louis, MO; 1 mg/kg) were administered via the jugular vein catheter. Injection volume was 1 ml. Measures of paw withdrawal latency (PWL) were taken at 10, 20, 30, 60, 90, and 120 min after saline or buprenorphine administration. A cut-off time of 15 s was set to prevent tissue damage of the hind paw.

Drug Administration and Recordings of Sleep/Wake States

A second series of experiments was designed to quantify the effect of intravenously administered buprenorphine on states of sleep and wakefulness. Buprenorphine was dissolved in sterile saline (pH 5.8 ± 0.2) and administered intravenously in a 1-ml volume at a dose of 1 mg/kg. Saline injection provided a negative control condition.

Recordings of sleep and wakefulness began at 08:00 at the initiation of the light phase of the light/dark cycle. Rats are nocturnal and light onset corresponds to the rat subjective night. In order to determine whether buprenorphine caused sleep disruption, as do other opioids,^26,37 this study was designed to deliver buprenorphine at light onset. Rats were placed in the recording chamber and the electromyogram and electroencephalogram electrodes were attached via swivel cable to the amplifiers and computer. All injections were administered during a 4-min interval. The data acquisition software was started when drug or vehicle administration began. The electroencephalogram signals were filtered between 0.3 and 30 Hz and amplified. Each rat (n = 7) received one injection of buprenorphine and one injection of saline separated by at least 1 week. The rats were then allowed to sleep and wake spontaneously for the remainder of the 24-h recording. At the end of the recording interval, rats were returned to the vivarium. Every 10 s of the 24-h recording was scored as wakefulness, nonrapid eye movement (NREM) sleep, or rapid eye movement (REM) sleep. All sleep recordings were also scored by a second individual who was blinded to the injection condition. There was a 93% agreement between the two sleep scorers.

A third series of experiments was designed to coadminister the nonbenzodiazepine sedative/hypnotic eszopiclone (Toronto Research Chemicals, Toronto, Canada) with buprenorphine in order to quantify the effect on sleep and wakefulness. Eszopiclone is a benzodiazepine receptor agonist with a non-benzodiazepine structure, marketed as Lunesta™ Eszopiclone is the (S)-isomer of the cyclopyrrolone zopiclone and is indicated for the treatment of insomnia.³⁸ As discussed in detail elsewhere,²⁶ a major complaint of patients who experience pain is poor sleep. Clinically used doses of opioids significantly disrupt sleep³⁷ and disordered sleep exacerbates pain.^26,39,40 These data raise the question of whether enhancement of sleep by a sedative/hypnotic would have a beneficial effect of diminishing opioid-induced sleep disruption. If so, this would encourage future studies aiming to determine if combining opioid and sedative/hypnotic therapy could diminish pain. Eszopiclone was dissolved in sterile saline and 1% dimethyl sulfoxide (pH 6.0 ± 0.2) and administered intravenously (3 mg/kg). Buprenorphine (1 mg/kg) was then delivered via the same intravenous cannula. For these studies, rats (n = 4) received an injection of eszopiclone followed immediately by an injection of saline or buprenorphine.

Measurement of Brain Adenosine Levels during Microdialysis Delivery of Buprenorphine

A fourth set of experiments sought to identify brain regions through which buprenorphine decreased sleep. Normal cholinergic neurotransmission is essential for maintaining wakefulness and opioids disrupt cholinergic neurotransmission in the SI region of the basal forebrain.³⁰ Adenosine is known to promote sleep and previous studies have shown that adenosine levels in the PnO are decreased by fentanyl and by morphine.²⁶ Both fentanyl and morphine cause sleep disruption. Therefore, the present experiments also used in vivo microdialysis and high performance liquid chromatography to measure adenosine levels in the PnO and SI during dialysis delivery of buprenorphine.

Buprenorphine (100 μM) was prepared the morning of each experiment. The drug was dissolved in Ringer's solution (pH 5.8 – 6.2) comprised of 146 nM NaCl, 4.0 mM KCl, 2.4 mM CaCl₂, and 10 μM erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA; Sigma-Aldrich) which is an adenosine deaminase inhibitor. Each rat was placed in an induction chamber and anesthetized with 4% isoflurane (Hospira, Inc.) in 100% oxygen. After 5 min, the rat was moved out of the chamber into a stereotaxic frame and fitted with a rat anesthesia mask, as described above. Isoflurane concentration was reduced to 2.5%. A midline scalp incision was then made to expose lambda and bregma. A Dremel tool (Racine, WI) was used to make a small craniotomy through which a dialysis probe could be placed in the brain. A rat brain atlas²⁹ was used to position a CMA-11 microdialysis probe (Cuprophane membrane: 1 mm long, 0.24 mm in diameter, 6-kDa cut off; CMA Microdialysis, North Chelmsford, MA) in the PnO or in the SI. Stereotaxic coordinates for the PnO were 8.4 mm posterior to bregma, 1.0 mm lateral to the midline, and 9.2 mm below bregma. The coordinates for the SI were 1.6 mm posterior to bregma, 2.5 mm lateral to the midline, and 8.7 mm below bregma. Delivered isoflurane concentration was held at 1.5% and was measured continuously throughout the duration of the experiment. A water blanket and recirculating heat pump (Gaymar Industries, Orchard Park, NY) were used to maintain body temperature at 37 degrees C throughout data collection and recovery.

The dialysis probe was perfused with Ringer's at a constant flow rate of 2 μl/min using a CMA/400 pump. Dialysis intervals of 15 min produced 30-μl samples which were injected into an high performance liquid chromatography system coupled to a UV-Vis detector (wavelength of 254 nm). This system made it possible to express measured adenosine levels as nM. The digitized chromatographs were quantified against a standard curve using ChromGraph software (Bioanalytical Systems). Adenosine levels were allowed to stabilize for 2 h before beginning data collection. A control sample was collected every 15 min for 1 h during dialysis with Ringer's. At the end of the 4th control sample, a liquid switch was activated to begin dialysis with Ringer's containing buprenorphine (100 μM). As noted elsewhere^26,41,42 the characteristics of the dialysis membrane are such that only about 5% of the 100 μM buprenorphine was delivered to the brain. After the final dialysis sample was collected, the probe was removed from the brain and the scalp incision was closed. The delivery of isoflurane was then discontinued and the animal was removed from the stereotaxic frame. The rats were returned to their cages and were monitored until they were ambulatory.

Histological Localization of Dialysis Sites

Four to five days after the microdialysis experiment each rat was deeply anesthetized and decapitated. Brains were removed, cut into 40-μm thick coronal sections with a cryostat (Leica Microsystems, Nussloch, Germany), and slide-mounted serially. Slides with brain sections containing the SI and PnO were fixed in paraformaldehyde vapor (80 degrees C) and stained with cresyl violet. All sections were then digitized using a Nikon Super Coolscan 4000 scanner (Tokyo, Japan). Coronal sections were compared to plates in a rat brain atlas²⁹ and the dialysis sites were localized to either the PnO or the SI.

Statistical Analysis

Statistical programs used for data analysis included Prism 5 (Graph Pad Software, Inc., La Jolla, CA) and SAS v9.2 (SAS Institute Inc., Cary, NC). All data were tested to insure they met the assumptions of the underlying statistical model. PWL was converted to Percent Maximum Possible Effect (% MPE) using the following equation: % MPE = (PWL experimental – PWL baseline)/(15 s – PWL baseline) × 100. Repeated measures, two-way analysis of variance (ANOVA) was used to analyze results for changes over time and changes due to drug, and Bonferonni post-hoc tests were used to detect differences at specific time points.

Every 10 s of the 24-h recordings of sleep and wakefulness was scored as wakefulness, NREM sleep, or REM sleep. Dependent measures included percent of time spent in each state, latency to onset of the first episode of NREM sleep and REM sleep, number of episodes, average episode duration, and number of transitions between states. To avoid the problem of inflated degrees of freedom resulting from the large number of 10-s epochs analyzed, the sleep-wake data were averaged for each rat. Dependent measures of sleep and wakefulness were analyzed by repeated measures two-way ANOVA and paired t-tests using Bonferonni correction.

As described in detail elsewhere^24,43,44 fast Fourier transform of the electroencephalogram was performed in order to determine whether the electroencephalogram was altered by buprenorphine. Electroencephalographic power was analyzed by repeated measures two-way ANOVA and post-hoc tests for comparison at every 0.5 Hz frequency band (Wake and REM sleep 5.0 to 10.0 Hz; NREM sleep 0.5 to 5.0 Hz).

For each experiment, adenosine measures during dialysis with Ringer's (control) were compared to adenosine levels during dialysis delivery of buprenorphine. This design allowed each experiment to contribute one mean adenosine value derived from four control (Ringer's) samples and one mean adenosine value derived from four measures obtained during administration of buprenorphine. These values were then averaged across multiple experiments and analyzed individually for PnO and SI brain regions using paired t-tests. A probability value of P ≤ 0.05 was considered to be statistically significant.

Results

Buprenorphine was Antinociceptive

Figure 1 depicts % MPE for paw withdrawal latency as a function of time after intravenous administration of saline and buprenorphine. ANOVA revealed that buprenorphine caused significant (P = 0.0072) antinociception. Bonferroni post-hoc comparisons indicated that buprenorphine significantly (P < 0.05) increased % MPE at 20, 30, 60, and 120 min after injection. This antinociceptive dose of buprenorphine was used for subsequent studies of sleep and wakefulness.

Figure 1

Buprenorphine increased latency of paw withdrawal (PWL) away from a thermal stimulus. Ordinate shows PWL expressed as percent maximum possible effect (% MPE). Abscissa shows time-course of % MPE relative to injection at time zero. Asterisks (*) indicate (more ...)

Buprenorphine Altered the Temporal Organization of Sleep and Wakefulness

Figure 2 illustrates the temporal distribution of wakefulness, NREM sleep, and REM sleep for 24 h following intravenous administration of saline (control) and buprenorphine. Figure 3 summarizes group data for the light phase (1^st 12 h after injection) showing buprenorphine-induced changes in the temporal organization of sleep and wakefulness. ANOVA indicated a significant (P < 0.01) effect of buprenorphine on percent of time spent in states of wakefulness, NREM sleep, and REM sleep, as well as a significant (P < 0.0001) drug-by-state interaction (fig. 3A). Paired t-tests with Bonferonni correction showed that buprenorphine significantly (P < 0.05) increased the percent of time spent in waking (25.2%) and significantly decreased the amount of time spent in NREM sleep (−22.1%) and REM sleep (−3.1%). Buprenorphine significantly delayed the onset of NREM sleep and REM sleep (fig. 3C).

Figure 2

Representative 24-h plots from one rat illustrating the buprenorphine-induced disruption of sleep architecture. Ordinate plots the temporal distribution of wakefulness (wake), nonrapid eye movement sleep (NREM), and rapid eye movement sleep (REM) for (more ...)

Figure 3

Buprenorphine altered the temporal organization of sleep and wakefulness and eszopiclone countered buprenorphine-induced sleep disruption. The left column summarizes the effect of buprenorphine relative to saline (control) on five dependent measures averaged (more ...)

There was a significant (P < 0.0001) drug main-effect and state-by-drug interaction (p < 0.0001) for the number of sleep/wake episodes (fig. 3E). Buprenorphine decreased the number of episodes of wakefulness (−88.2%), NREM sleep (−89.5%), and REM sleep (−90.8%). Figure 3G shows that buprenorphine significantly (P < 0.0001) altered the duration of sleep/wake episodes. Average duration of wakefulness was significantly increased (529.6%) and the duration of sleep epochs was decreased for both NREM sleep (−30.8%) and REM sleep (−87.5%). Figure 3I shows that buprenorphine also significantly (P < 0.0001) decreased the number of transitions (−89.8%) between states.

Figure 4 plots the percent state for each drug condition during the 12-h dark phase (rat subjective day) of the light/dark cycle that followed the 12-h light phase depicted by figure 3. Within the dark phase, when rats are normally awake and active, the time spent awake was significantly (P = 0.012) decreased by buprenorphine. The buprenorphine condition within the dark phase revealed significantly (P = 0.0019) more NREM sleep and a nonsignificant decrease in REM sleep compared to the saline condition.

Figure 4

The percent of time during the 12-h dark phase that rats (n = 7) spent in states of wakefulness (Wake), nonrapid eye movement sleep (NREM), and rapid eye movement sleep (REM). These recordings were obtained during hours 12 through 24 after administration (more ...)

The effect of buprenorphine on states of sleep and wakefulness can also be visualized by comparing the light phase (fig. 3A) and dark phase (fig. 4) results. NREM sleep after buprenorphine increased significantly (P = 0.0003) from an average of 5.5% in the light phase (fig. 3A) to 27.4% in the dark phase (fig. 4). There was also a significant (P = 0.003) rebound increase in REM sleep from an average of 0.33% after buprenorphine during the light phase (fig. 3A) to about 4% after buprenorphine during the dark phase (fig. 4).

Eszopiclone Decreased the Sleep Disruption Caused by Buprenorphine

The five illustrations in the right column of figure 3 summarize the results of experiments designed to determine whether the sedative/hypnotic eszopiclone countered the buprenorphine-induced inhibition of sleep. Eszopiclone when coadministered with buprenorphine prevented the significant increase in wakefulness (fig. 3B) caused by buprenorphine alone (fig. 3A). Similarly, the significant buprenorphine-induced decrease in NREM sleep and REM sleep (fig. 3A) was prevented by coadministration of eszopiclone (fig. 3B). Eszopiclone blocked the significant increase in latency to sleep onset (figs. 3C vs. 3D). Eszopiclone partially reversed the buprenorphine-induced decrease in both the number of wakefulness and NREM sleep episodes (fig. 3E vs. 3F). The 530% increase in average duration of waking episodes caused by buprenorphine (fig. 3G) was reduced to a 171% increase by coadministration of eszopiclone (fig. 3H). Eszopiclone blocked the significant decrease in number of state transitions caused by buprenorphine (fig. 3I vs. 3J).

Buprenorphine Increased Electroencephalogram Delta Power during NREM Sleep

Figures 5A-C illustrate electroencephalogram power recorded across states of sleep and wakefulness after intravenous administration of buprenorphine to awake, freely moving rats. Buprenorphine did not alter electroencephalogram power during wakefulness or REM sleep (figs. 5A & C), but did increase electroencephalogram power in the delta frequency range during NREM sleep (fig. 5B). ANOVA revealed a significant (P = 0.007) buprenorphine main-effect on electroencephalogram frequency bands ranging from 0.5 to 5.0 Hz in 0.5 Hz increments (fig. 5B). The fast Fourier transform analyses were conducted for electroencephalogram measures obtained during the 12-h light period (i.e., rat's subjective night) that immediately followed buprenorphine administration. As figures 2 and and33 show, buprenorphine depressed NREM sleep for 6 to 8 h. Measurement of the increase in electroencephalogram delta power was conducted for up to 12-h after buprenorphine administration. A future study will be needed to determine whether, and for how long beyond 12-h, electroencephalogram delta power is increased by buprenorphine.

Figure 5

Electroencephalographic power (ordinate) plotted as a function of frequency (abscissa) averaged for 5 rats across states of sleep and wakefulness. During wakefulness (A) and rapid eye movement (REM) sleep (C) buprenorphine did not change electroencephalographic (more ...)

Buprenorphine Decreased Adenosine Levels in PnO and SI

Histological analyses confirmed that all microdialysis sites were localized to the PnO or to the SI (fig. 6A). Figure 6B shows the results of one representative experiment. Adenosine levels in the SI are plotted as a function of time during dialysis with Ringer's (121-180 min after probe placement) followed by dialysis delivery of buprenorphine (181-240 min after probe placement). Figures 6C and D confirm chromatographic identification of adenosine Figure 6C illustrates chromatograms produced by five known concentrations of adenosine. Figure 6D shows chromatograms reflecting brain adenosine (dialyzed Ringer's), a negative control (nondialyzed Ringer's), a positive control (brain adenosine sampled during dialysis delivery of the adenosine deaminase inhibitor EHNA), and an adenosine standard.

Figure 6

Microdialysis delivery of buprenorphine decreased adenosine levels in the basal forebrain and pontine reticular formation. Cresyl-violet stained coronal sections show representative dialysis sites in (A) the pontine reticular formation, oral part (PnO) (more ...)

Figure 7A summarizes a final set of experiments that quantified adenosine levels in SI and PnO as a function of route of buprenorphine administration. Microdialysis delivery of buprenorphine significantly (P = 0.03) decreased adenosine levels in PnO (−14.8%) and significantly (P = 0.0004) decreased adenosine levels in the SI region of the basal forebrain (−36.7%). Figure 7B plots adenosine levels in the SI before and after intravenous administration of buprenorphine to isoflurane-anesthetized rat. Buprenorphine significantly (P < 0.0001) decreased (−20.3%) adenosine levels in the SI.

Figure 7

Central and systemic delivery of buprenorphine decreased brain adenosine levels. (A) Microdialysis delivery of buprenorphine to the pontine reticular formation, oral part (PnO) (n=5 rats) or to the substantia innominata (SI) (n=5 rats) decreased adenosine (more ...)

Discussion

Buprenorphine can be efficacious in the treatment of opioid and heroin addiction,^45-47 and there is increasing interest in the use of buprenorphine for pain management.^27,28 The analgesic effects of buprenorphine are mediated, in part, via agonist actions at the μ opioid receptor.⁴⁸ This is the first study presenting electrographic data that demonstrate significant sleep disturbance (fig. 3) caused by an antinociceptive dose of buprenorphine (fig. 1). The present finding that an antinociceptive dose of buprenorphine disrupts sleep is discussed relative to the relationship between sleep and nociception, the potential for developing counter-measures for opioid-induced sleep disruption, and the underlying mechanisms.

Buprenorphine Disrupted Sleep

Some data suggest that buprenorphine is superior to traditional opioids for the treatment of pain due to its reported analgesic and antihyperalgesic effects with fewer side effects (low incidence of respiratory depression and less constipation).²⁸ Buprenorphine shares some similar pharmacodynamic properties with traditional opioids, and patient-report data indicate benefits from buprenorphine therapy. Freye et al.²⁷ found that self-report sleep quality rated as “good” or “very good” increased from 14% to 74% when patients were transitioned from high-dose oral morphine to transdermal buprenorphine. Transdermal buprenorphine has been compared to placebo for ability to decrease pain and promote sleep, and patients randomized to receive buprenorphine report less pain and improved sleep.⁴⁹ Specifically, subjects who received buprenorphine reported less trouble falling asleep, decreased requirement for sleeping medication, and decreased awakening at night caused by pain. Another study found a nonsignificant trend of improved sleep favoring transdermal buprenorphine over extended release tramadol tablets for the treatment of osteoarthritis.⁵⁰ A known limitation of such studies is that self-assessment of sleep quality may not show faithful concordance with objective, electrographic measures of sleep.⁵¹

There is a growing appreciation for the interrelationship between sleep and pain.³⁹ Sleep deprivation in healthy normals lowers pain perception thresholds.⁵² The chronic effects of μ-opioid receptor agonists on sleep in pain patients are not completely understood.³⁹ Opioids cause sleep disturbance^4,37,53 and the present results demonstrate that buprenorphine increases wakefulness and disrupts the temporal organization of sleep (figs. 2--4).4). Sleep, like breathing, is an endogenously generated biological rhythm. Just as rhythmic switching from inspiration to expiration is essential for gas exchange, the ability of sleep to produce reports of rest and recovery requires a normal temporal organization. Buprenorphine caused a decrease in the number (fig. 3E) and an increase in the duration (fig. 3G) of wakefulness episodes. The decreased number of state transitions (fig. 3I) reflects the buprenorphine-induced disruption of sleep continuity. Figure 4 summarizes the percentage of time spent in states of sleep and wakefulness during the 12-h dark phase when rats are normally active. These dark phase recordings were continuous with the figure 3 data during the 12-h light phase. Thus, the figure 4 data show that for 12 to 24 h after administration of buprenorphine there was a rebound increase in sleep at a time when nocturnal rodents are normally most active. The potential clinical relevance of buprenorphine-induced disruption of sleep continuity derives from the fact that repeated sleep disruption negatively impacts neurocognitive function as severely as does total sleep deprivation.⁵⁴

Opioid-induced sleep disruption has the potential to negatively impact patient care because sleep deprivation is known to lower pain threshold.^15,16,52 This study did not address the impact of pain or the treatment of pain on sleep disturbance. Some believe that medications from the agonist/antagonist class, such as buprenorphine, may be less associated with the adverse effects of traditional μ-agonists; however, the present data indicate that the sleep-disrupting effects of buprenorphine are similar to those of other opioids.^26,30,37,53 The present results encourage studies designed to objectively quantify the effects of buprenorphine on sleep in humans.

The U.S. Food and Drug Administration approved buprenorphine for the treatment of opioid addiction. Suboxone (buprenorphine/naloxone sublingual tablet, Reckitt Benckiser Pharmaceuticals Inc., Richmond, VA), Subutex (buprenorphine sublingual tablet, Reckitt Benckiser Pharmaceuticals Inc.), and transdermal buprenorphine (not available in the United States) are increasingly used to treat pain. The present finding of buprenorphine-induced sleep disruption also is relevant to evidence indicating that sleep disturbance can lead to a higher potential for addiction relapse.^55-57

Electroencephalogram delta waves (0.5 to 4 Hz) are one of three rhythmic waveforms characteristic of NREM sleep.⁵⁸ Delta waves provide an index of sleep intensity^59,60 and electroencephalogram power in the delta frequency increases during recovery sleep that follows sleep deprivation.^61,62 The finding that buprenorphine caused an increase in electroencephalogram delta power during NREM sleep (fig. 5B) is consistent with subjective reports that buprenorphine improves sleep^7,27 and with evidence that opioids increase delta power in humans.⁶³ A search of the Medline database from 2001 to 2011 revealed no polysomnographic data characterizing the effect of buprenorphine on human sleep.

Eszopiclone was an Effective Counter-Measure that Prevented Sleep Disruption by Buprenorphine

Buprenorphine significantly disrupted the temporal organization of sleep (fig. 3, left column). Most aspects of sleep disturbance were prevented by cotreatment with the nonbenzodiazepine hypnotic eszopiclone (fig. 3, right column). Sedative/hypnotics are a standard treatment for insomnia, but their effects in the treatment of pain- and opioid-induced sleep disturbance are still poorly understood. The use of sedative/hypnotics may not be a mainstay of addiction therapy, but the present results (fig. 3, right column) indicate their potential to prevent buprenorphine-induced sleep disturbance. Eszopiclone as an adjunctive agent coadministered with the antidepressant fluoxetine resulted in a faster onset and greater magnitude of the desired antidepressant effect.⁶⁴ An exciting area open to future study is to determine whether hypnotics can be used as an effective counter-measure for opioid-induced sleep disruption.

Limitations, Potential Clinical Relevance, and Conclusions

The present study was designed to quantify the effects of burprenorphine on sleep and adenosine levels. The results are limited to documenting that burprenorphine, similar to morphine and fentanyl, disrupted sleep and decreased adenosine levels in sleep-related brain regions. The results do not imply that the effects of burprenorphine were mediated only by μ opioid receptors. Burprenorphine may have disrupted sleep and decreased adenosine, in part, by acting as a κ antagonist.

The analgesic²¹ and sleep-promoting⁶⁵ effects of adenosine are well known and suggest adenosine as a molecule of potential clinical relevance for anesthesiology. There is good agreement between preclinical and clinical data that opioids disrupt sleep³⁷, a finding confirmed by administering opioids to pain free humans.⁵³ The restorative effects of sleep require normal temporal organization of sleep. Unfortunately, morphine and fentanyl slow the electroencephalogram during wakefulness, increase lighter stage 2 NREM sleep, decrease stage 3 and 4 NREM sleep, and decrease or eliminate REM sleep. Disruption of normal sleep impairs immune function, exacerbates pain³⁹, and, particularly in older patients, can be a precipitating factor for postoperative delirium.⁶⁶

In conclusion, the results show for the first time that buprenorphine disrupted normal sleep architecture and decreased adenosine levels in sleep regulating regions of the basal forebrain and pontine reticular formation (figs. 6 and and7).7). The buprenorphine results are consistent with the discovery that fentanyl and morphine decrease adenosine levels in basal forebrain and pontine reticular formation.²⁶ The present study extends the earlier findings by providing mechanistic insights into a brain site and a molecule by which buprenorphine disrupts sleep. Novel insights were obtained by holding site of adenosine measurement constant within the substantia innominata region of the basal forebrain while varying route of buprenorphine delivery. The results show that both microdialysis delivery to the substantia innominata and systemic administration of buprenorphine caused a significant decrease in adenosine in the substantia innominata. As demonstrated elsewhere^30,44 when effects caused by drug delivery to a specific brain region replicate the effects caused by systemic delivery, it is logical to infer that the actions of systemically administered drugs are mediated, in part, by that brain region and by the neurotransmitter molecule being measured. Thus, the neurochemical results, combined with the sleep disrupting effect of buprenorphine, support the interpretation that one mechanism through which buprenorphine disrupts sleep is by decreasing adenosine levels in the substantia innominata region of the basal forebrain.

Acknowledgments

For expert assistance the authors thank Sha Jiang, B.S.m Research Associate, Mary A. Norat, B.S., Senior Research Associate, and Sarah L. Watson, B.S., Senior Research Associate, from the Department of Anesthesiology, and Kathy Welch, M.A., M.P.H., Statistician Staff Specialist, Center for Statistical Consultation and Research, University of Michigan, Ann Arbor, Michigan.

Support: Supported by grants HL40881 (RL), HL65272 (RL), and MH45361 (HAB) from the National Institutes of Health, UL1RR024986 (CMB) from the National Center for Research Resources, Bethesda, Maryland, and by the Department of Anesthesiology, University of Michigan, Ann Arbor, Michigan

Footnotes

Attribution: Department of Anesthesiology, University of Michigan, Ann Arbor, Michigan

Meetings at which work was presented: Abstracts presented at the American Society of Pain Meeting, Baltimore, Maryland, May 6, 2010.

^*http://www.nap.edu/catalog.php?record_id=12910. Last accessed May 31, 2011.

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