European Journal of Pharmacology 459 (2003) 217 – 219
`
`www.elsevier.com/locate/ejphar
`
`Short communication
`
`Naloxone displacement at opioid receptor sites measured in vivo in the
`human brain
`
`Jan K. Melichar, David J. Nutt*, Andrea L. Malizia
`
`Psychopharmacology Unit, School of Medical Sciences, University of Bristol, University Walk, Bristol BS8 1TD, UK
`
`Received 12 November 2002; received in revised form 2 December 2002; accepted 6 December 2002
`
`Abstract
`
`We report the use of a sensitive non-tomographic positron detecting system to measure the dose – response curve of naloxone in human
`brain. [11C]Diprenorphine was administered to normal volunteers in tracer amounts and, 30 min later, various bolus doses of naloxone were
`given (1.5 – 160 Ag/kg) intravenously and change in [11C]diprenorphine binding monitored over the next 30 min. We found that this method
`produced results consistent with existing data. It was observed that f13 Ag/kg of naloxone (f1 mg in an 80 kg man) was required to produce
`an estimated 50% receptor occupation. This is consistent with the clinical dose of naloxone used to reverse opiate overdose (0.4 mg – 1.2 mg).
`D 2002 Elsevier Science B.V. All rights reserved.
`
`Keywords: Opioid receptor; Opioid receptor antagonist; Diprenorphine; Naloxone; PET (Positron-Emission Tomography)
`
`1. Introduction
`
`Drugs active at the opioid receptor are important in a
`variety of clinical conditions, as well as in the field of
`addiction. Measures of opioid receptor binding and occu-
`pancy in man in vivo can be obtained using [11C]diprenor-
`phine, a positron-emitting ligand. This has been used with
`Positron-Emission Tomography (PET) to delineate the dis-
`tribution of opioid receptors in the human brain (Jones et al.,
`1988) and in the study of a variety of clinical conditions,
`including epilepsy (Bartenstein et al., 1993) and pain (Jones
`et al., 1999). [11C]Diprenorphine is a weak partial opiate
`agonist which is structurally similar to naloxone (a full opiate
`antagonist) and labels A, n and y brain opioid receptors with
`similar affinities (Jones et al., 1988; Sadzot et al., 1991;
`Seeman, 1993). In conventional PET scans, it takes 20 – 30
`min for [11C]diprenorphine to reach maximal levels in the
`brain, with a stable level of activity or slow decline being
`observed in opioid receptor-rich regions (e.g. frontal, tempo-
`ral and parietal cortices) in the following 30 min.
`The Multiple Organs Coincidences Counter (MOCC) is a
`non-tomographic alternative to PET. It is a whole body gam-
`ma-ray counter modified to detect coincident counts from
`
`* Corresponding author. Tel.: +44-117-925-3066; fax: +44-117-927-
`7057.
`E-mail address: david.j.nutt@bristol.ac.uk (D.J. Nutt).
`
`whole regions of the body and is very sensitive, detecting
`radiolabelled tracers given at <1% of the dose used in PET.
`The modification measures coincident counts between pairs
`of highly sensitive sodium iodide detectors and gives a field
`of view of approximately 10 cm. Thus, the left chest is used,
`where necessary, as a reference region, as it is not possible to
`delineate separate areas of the brain with this instrument.
`Reproducible time activity curves of exactly the same tem-
`poral profile to those from conventional PET can be obtained
`from whole body areas exposing patients to 50 – 100 ASv of
`radiation as opposed to 1.5 – 2.5 mSv with conventional PET
`(Malizia et al., 1995).
`Naloxone is a full opiate antagonist and is used clinically
`for the complete or partial reversal of narcotic depression,
`including respiratory depression, induced by opioids includ-
`ing natural and synthetic narcotics. It is also indicated for the
`diagnosis of suspected acute opioid overdosage, where an
`initial dose of 0.4 mg to 1.2 mg is usually administered
`intravenously (occasionally up to 10 mg is given).
`This study aims to validate the use of pulse chase (or
`displacement) experiments in the MOCC to measure nalox-
`one occupation in the human brain and to assess the relation-
`ship between naloxone occupation and the doses required in
`clinical use. Previous work in the MOCC has validated this
`paradigm using the tracer flumazenil to measure GABAA
`receptor sites (Malizia et al., 1995). Villemagne et al. (1994)
`have previously conducted studies on the opioid receptors
`
`0014-2999/02/$ - see front matter D 2002 Elsevier Science B.V. All rights reserved.
`doi:10.1016/S0014-2999(02)02872-8
`
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`218
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`J.K. Melichar et al. / European Journal of Pharmacology 459 (2003) 217–219
`
`using a similar instrument, though these studies used an
`equilibrium ‘preloading’ paradigm. The advantage of the
`‘displacement’ paradigm used in the present study is that all
`the data can be collected in a single experiment. Calculation
`of occupancy measures may be biased by changes in tissue
`perfusion. Naloxone has been reported in particular condi-
`tions to change brain blood flow (Nagamachi et al., 1995;
`Komjati et al., 2001).
`
`2. Materials and methods
`
`Five healthy volunteers were studied in the MOCC,
`receiving an initial injection of f80 ACi of [11C]diprenor-
`phine. The activity was recorded from the head and the left
`chest for 1 h after injection (the maximum possible, given the
`short half life of [11C]diprenorphine and the low radiation
`doses given), and analysed as previously described (Malizia
`et al., 1995; Melichar et al., 2001). Thirty minutes after the
`initial injection, they received a bolus i.v. injection of
`naloxone and the change in gradient of the washout curve
`before and after administration of naloxone was measured.
`Three volunteers had only one study, one had a further two
`studies and another had a further three studies (in these two
`individuals, they received different doses of naloxone in each
`study and in one of their studies received no naloxone).
`Thus, in total, two control studies and eight naloxone dis-
`placement studies were done. Doses of naloxone given were
`1.5, 4, 5, 10, 12.5, 15, 80 and 160 Ag/kg. For an 80-kg man,
`this corresponds to 0.12, 0.32, 0.4, 0.8, 1, 1.2, 6.4 and 12.8
`mg in total, respectively. The protocol was approved by the
`Administration of Radioactive Substances Advisory Com-
`mittee (ARSAC) and the local Ethics Committee.
`
`3. Results
`
`The signals from the head were much greater than those
`from the left chest and demonstrated an increased [11C]dipre-
`
`Fig. 1. Typical [11C]diprenorphine MOCC study, illustrating significant
`reduction of signal when naloxone bolus given.
`
`Fig. 2. Log dose – response curve: naloxone displacement of [11C]diprenor-
`phine in the MOCC in eight healthy volunteers—gradient changes.
`
`norphine washout rate after injection of naloxone (see
`Fig. 1), which continued until the end of the study (a further
`30 min). This change in washout rate was measured and the
`results are shown in Fig. 2 in the traditional log dose –
`response format. From this, it is estimated that 50% of opioid
`receptors in the brain are occupied by naloxone at a dose of
`approximately 13 Ag/kg.
`
`4. Discussion
`
`These results demonstrate that this method is useful for
`calculating opioid receptor occupation in man in vivo. We
`have shown that the dose needed to occupy 50% of available
`receptors in the adult human brain is approximately 13 Ag/
`kg, which, in an 80-kg man, corresponds to 1.04 mg. This
`closely mirrors the doses given in clinical practice for the
`treatment of opioid overdose (usually 0.4 to 1.2 mg, i.e. one
`to three ampoules at 0.4 mg/ampoule). These data are
`consistent with previous observations by Villemagne et al.
`(1994) who used a preloading paradigm. We can therefore
`conclude that a displacement paradigm seems effective in
`healthy volunteers. It can also be inferred that to block the
`effects of an overdose of opioid agonists with naloxone
`requires around 50% blockade of available opioid receptors,
`given the results from this study and everyday clinical
`practice with naloxone.
`However, this study does not tell us what occurs in the
`brains of opiate-dependent individuals—further experiments
`will be needed before extrapolating these results to this
`group. This is because these individuals may well have
`changes in receptor numbers due to their chronic misuse of
`opiates and might exhibit large changes in cerebral blood
`flow when exposed to naloxone (Zamani et al., 2000).
`In summary, this is a valid technique for deriving an index
`of opioid receptor site occupation using very low doses of
`radiation which allows repeated measurements to be made.
`We have shown that this method estimates 50% receptor
`occupation by f13 mg/kg of naloxone in man in vivo,
`which is in line with both pre-clinical work and clinical
`experience.
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`219
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`Acknowledgements
`
`We are grateful for the help and support of the staff at the
`MRC Cyclotron Centre in London, especially Dr. Roger
`Gunn, as well as Dr. Judy Myles, Dr. Anne Lingford-Hughes
`and Dr. Mark Daglish in Bristol and to the Wellcome Trust
`and the Medical Research Council for the financial support.
`
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