Opioid µ, δ and κ Receptors for Endorphins

8 Pages·2000·0.09 MB·English

by  Carr J.J., Blaloek J.E.

Opioid (cid:22), (cid:14), and
(cid:20) Receptors for
Endorphins
Daniel J. J. Carr1 and J. Edwin Blalock2,*
1Department of Microbiology and Immunology, LSU Medical Center, New Orleans,
LA 70112-1393, USA
2Department of Physiology and Biophysics, University of Alabama at Birmingham,
Birmingham, AL 35294-0005, USA
*corresponding author tel: [phone], fax: [phone], e-mail: [email protected]
DOI: 10.1006/rwcy.2000.23005.
SUMMARY ((cid:11),(cid:12),and(cid:13))(Bradburyetal.,1976;Coxetal.,1976),
dynorphin (Goldstein et al., 1979), and endomorphin
(Zadina et al., 1997), suggested the existence of
Thethreemajortypesofopioidreceptors((cid:22),(cid:14),and(cid:20))
multiple types of receptors (termed opioid receptors)
are members of the DRY (Asp-Arg-Tyr)-containing
for these natural ligands. Evidence for multiple types
subfamily of seven transmembrane spanning recep-
of opioid receptors was obtained using congeners of
tors. Opioid receptors on cells of the immune system
morphineinspinalstudiesindogs(GilbertandMartin,
are virtually identical to those on neuronal cells. The
1976; Martin et al., 1976).
activation of opioid receptors in the CNS leads to
Originally identified in the early 1970s (Pert and
analgesia while activation of opioid receptors on
Snyder, 1973; Simon et al., 1973; Terenius, 1973),
immune cells can enhance or suppress immune func-
today,threemaintypesofopioidreceptorshavebeen
tiondependingonthetarget cellandimmuneparam-
defined and cloned: (cid:14), (cid:20), and (cid:22) opioid receptors with
eter measured.
pharmacologically distinct subtypes for (cid:14) ((cid:14) and (cid:14) ),
1 2
(cid:20) ((cid:20) , (cid:20) , and (cid:20) ), and (cid:22) ((cid:22) and (cid:22) ) (Pasternak,
1 2 3 1 2
1993).Twootherreceptorsincluding"(specificfor(cid:12)-
BACKGROUND
endorphin) and (cid:27) receptors were originally described
as opioid receptors but have since been redefined as
Discovery
nonopioid (Simon, 1991). All of these receptors have
been identified and characterized on cells of the
The molecular characterization of opioid receptors immune system (Garza and Carr, 1997).
has been investigated for nearly 25 years. However, Another opioid-like receptor, referred to as the
the activities of these receptors, as manifested in the nociceptivereceptor(orphanopioidreceptor),origin-
effectsofopioidcompounds(e.g.opium,ofwhichthe ally described in human brainstem (Mollereau et al.,
mainactiveingredientismorphine),havebeenknown 1994) was also found in mouse spleen T and B lym-
foratleast6000years,sincethetimeoftheSumerians phocytes,whereitwasfirstcoupledtoaphysiological
(4000 BC). The discovery of endogenous opioid pep- role (Halford et al., 1995). This review focuses on the
tides, including the enkephalins (met- and leu- immunecell-derivedopioidreceptors,comparingtheir
enkephalin) (Hughes et al., 1975), the endorphins physicochemical properties with those found in the
2212 Daniel J. J. Carr and J. Edwin Blalock
nervous system as well as defining their role in the example, (cid:20) opioid receptors bound to (cid:20)-selective
immune system. opioidligandshavebeenfoundtoreducesignificantly
monocytotropic HIV-1 SF162 strain replication in
microglia-enriched cultures (Chao et al., 1996).
Structure
GENE
The three major types of opioid receptors are mem-
bers of the (DRY)-containing subfamily of seven
transmembrane spanning receptors. There is 60% Accession numbers
aminoacididentitybetweeneachtypeofopioidrecep-
tor with the membrane-spanning regions (transmem- The (cid:14) (L06322, L11065), (cid:20) (L11064), and (cid:22) (L22455,
brane I–VII) and the intracellular loops connecting L20684)opioidreceptorshavebeen clonedfrom neu-
thesesegmentsbeinghighlyconservedbetweenrecep- ronal tissue (Evans et al., 1992; Kieffer et al., 1992;
tor types. Studies indicate that ligands (agonists and Li et al., 1993; Thompson et al., 1993; Wang et al.,
antagonists) to these receptors bind to different 1993; Yasuda et al., 1993).
regions of the extracellular domain and such inter-
action can be greatly influenced by the transmem-
brane segments (predominantly TM II, III, and VI) PROTEIN
(Kong et al., 1993, 1994; Surratt et al., 1994). Also,
changes in one amino acid in the TM IV spanning
Accession numbers
region has been shown to alter opioid antagonist to
agonist activity (Claude et al., 1996). Since the amino
Protein Information Resource:
acid sequences of the neuronal- and some immune-
Human (cid:22) opioid receptor: 2135858
derived receptors are nearly identical, it is predicted
Human (cid:20) opioid receptor: 631277
that a similar relationship between agonist/antago-
Human (cid:14) opioid receptor: 2134989
nist-binding domains and the influence of the trans-
membrane spanning regions will be found in the
immune-derivedopioidreceptors.However,immune-
Sequence
derived opioid receptor-binding domains according
to some investigators may be distinct since binding
The neuronal opioid receptors are composed of
or biochemical characteristics of these sites are not
between370and389aminoacidsencodedbymRNAs
characteristic of neuronal opioid sites (e.g. Stefano
ranging in size from 1.9 to >10.0kb (Carr et al.,
et al., 1992; Makman et al., 1995).
1996). Both (cid:14) (Figure 1a; Sedqi et al., 1996) and (cid:20)
(Figure1b;Belkowskietal.,1995)receptorfull-length
Main activities and cDNAs (predicted to be 372–400 amino acids in
length) have been identified in thymocytes or a
pathophysiological roles
thymoma cell line. However, only a partial sequence
(441bp) of a (cid:22) opioid receptor has been identified by
Theprimaryfunctionassociatedwithneuronalopioid RT-PCR in peripheral blood mononuclear cells
receptorsis thecontrolofthe sensationofpaineither (Chuanget al.,1995) andrat peritonealmacrophages
through receptors located spinally ((cid:14) , (cid:20) , and (cid:22) ) or (Figure 1c; 721bp) (Sedqi et al., 1995). All immune
1 1 2
supraspinally ((cid:14) , (cid:20) , (cid:20) , and (cid:22) ) (Pasternak, 1993). cell-derived receptor sequences identified thus far are
2 2 3 1
Withintheimmunesystem,opioidreceptorsfoundon nearlyidentical((cid:21)99%homology)withthereceptors
immunecellsmayaugmentorsuppressimmunefunc- in the nervous system.
tiondependingonthecelltypeandstimulation(Carr,
1991). However, alkaloid opioid ligands (e.g. mor-
Description of protein
phine and fentanyl) are potent immunosuppressive
compoundsaffectingtheimmunesystemprimarilyby
indirect pathways ligating to receptors found within Byavarietyoftechniques,theneuronalopioidrecep-
the CNS and activating secondary systems (including torswereobservedtorangeinsizefrom40to65kDa
the adrenergic pathway and the hypothalamus- (Simonds, 1988; Loh and Smith, 1990; Wollemann,
pituitary-adrenalaxis)(Carretal.,1996).Otherfunc- 1990). The data concerning the biochemical proper-
tionsofimmune-derivedopioidreceptorsmaypertain tiesofthesereceptorsmaybelimitedbytheuncertain
to the response to infectious pathogens. As an specificities of some of the antireceptor antisera.
Opioid (cid:22), (cid:14), and (cid:20) Receptors for Endorphins 2213
Figure1 (a)Deducedaminoacidsequence labeledproteinsmigratingat70,46,and31kDafrom
of the (cid:14) opioid receptor cloned from brain tissue, while a major protein species migrating
activated murine thymocytes as reported at 31kDa was labeled from spleen tissue. The31kDa
by Sedqi et al. (1996). Bold letters indicate species was thought to be a degradative form of the
changes from the published rodent brain (cid:14)
mature protein. Subsequent analysis of immune cell-
opioid receptor. (b) Deduced amino acid
derived (cid:14), (cid:20), and (cid:22) opioid receptors determined the
sequence of the (cid:20) opioid receptor cloned
size to be nearly identical to that of the neuronal
fromR1.1thymomacelllineasreportedby
receptors (Carr, 1991).
Alicea et al. (1998). Bold letters indicate
changes from the published rodent brain (cid:20)
opioidreceptor.(c)Deducedaminoacidse-
quenceofthe(cid:22)opioidreceptorclonedfrom Relevant homologies and species
adherentperitonealmacrophagesasreported
differences
by Sedqi et al. (1995). Bold letters indicate
changes from the published rodent brain (cid:22)
opioid receptor. Opioid receptors from immune cells are virtually
identical ((cid:21)99% homology) to those on neuronal
(a) cells.Thevariousopioidreceptortypes((cid:22),(cid:14),(cid:20))show
about 60% homology.
MELVPSARAELQSSPLVNLSDAFPSAFPSA 30
GANASGSPGARSASSLALAIAITVLYSAVC 60
AVGLLGNVLVMFGIVRYTKLKTATNIYIFN 90
LALADALATSTIPFQSAKYLMETWPFGELL 120 Affinity for ligand(s)
CKAVLSIDYYNMFTSIFTLTMMSVDRYIAV 150
CHPVKALDFRTPAKAKLIQICIWVLASGVG 180
VPIMVMAVTQPRDGAVVCMLQFPSPSWYWD 210 Theidentificationofthetypesofopioidreceptorshas
TVTKICVFLFAFVVPILIITVCYGLMLLRL 240 been greatly facilitated by the design and synthesis of
RSVRLLSGSKEKDRSLRRITRMVLVVVGAF 270 opioid ligands selective for the types of receptors to
VVCWAPIHIFVIVWTLVDINRRDPLVVAAL 300
which they bind (Table 1). Similar to cell-associated
HLCIALGYANSSLNPVLYAFLDENFKRCFR 330
neuronalopioidreceptors,theclonedneuronalopioid
QLCRTPCGRQEPGSLRRPRQATTRERVTAC 360
TPSDGPGGGAAA 372 receptors expressed in PC-12 cells showed high-
affinitybindingtoligandsrangingfrom0.2to3.0nM
(b) (Raynoret al.,1994).Opioidreceptorsfoundoncells
of the immune system display a modestly reduced
MESPIQIFRGNPGPTCSPSACLLPDSSSWF 30
affinity for their ligands ranging from 20 to 900nM,
PDWAESNSDGSVGSENQQLESAHISPAIPV 60
IITAVNSVVFVVGLVGDSLVMFVIIRIYTK 90 depending on the ligand and receptor (Garza and
MKTATDIYIFDLALANALVTTTMPFQSAVY 120 Carr, 1997). For example, (cid:20) receptors display affin-
LMDSWPFGNVLCKIVISINYYDMFTSIFTL 150 ities ranging from 4.1 to 65.0nM (Garza and Carr,
TMMSVNRYIAVCHPVKALNFRTPLKAKIID 180
1997), whereas a unique alkaloid-specific (cid:22) opioid
ICIWLLASSVGISAIVLGGTKVRENVNVIE 210 3
CSLQFPNNEYSWWNLFMKICVFVFAFVIPV 240 receptor found on granulocytes has a Kd of 44nM
LIIIVCYTLMILRLKSVRVLSGSREKNRDL 260 (Makmanetal.,1995).Oneinvestigationreportedthe
RRITKLVLVVVAVFIICWTPIHIFILVEAL 290 IC for an immunoaffinity-purified opioid receptor
50
GSTSHSTAALSSYYFCIALGYTDSSLDPVL 320 isolated from mouse spleen preparations to be
YAFLNEDFKRCFRNFCFPIKMRMERQSTDR 350
approximately 700nM, suggesting a loss in affinity
DTVQNPASMRNVGGMDKPV 369
upon purification (Carr et al., 1990).
(c)
MGTWPFGTILCKIVISIDYYNMFTSIFTLC 30 Cell types and tissues expressing
TMSVDRYIAVCHPVKALDFRTPRNAKIVNV 60
CNWILSSAIGLPVMFMATTKYRQGSIDCTL 90 the receptor
TFSHPTWYWQNLLKICVFIFAFIMPILIIT 120
VCYALMILRLKSVRMLSGSKEKNRDLRRITR 150
Within the immune system, there is some disagree-
MVLVVVAVFIVCWTPIHIYVIIKALITIPE 180
TTFQTVSWHFCIALGYTDSCLDPVLYAFLN 210 ment as to the population of cells that express opioid
receptors.Tothisend,Table2andTable3summarize
Using a site-directed acylating agent derived from the evidence for the presence of opioid receptors on
fentanyl(knownassuperfit)thatishighlyselectivefor primary cells of the immune system (Table 2) and
(cid:14) opioid receptors, a comparison of the mouse brain cell lines derived from cells of the immune system
cell-andspleencell-derived(cid:14)opioidreceptor.Superfit (Table 3) based on pharmacological (radioreceptor
2214 Daniel J. J. Carr and J. Edwin Blalock
Table 1 Commercially available selective opioid agonists/antagonists
(cid:14) Opioid receptor ligands (cid:20) Opioid receptor ligands (cid:22) Opioid receptor ligands
DADLE (agonist) Bremazocine (agonist) DAMGO (agonist)
DPDPE (agonist) U-50488 (agonist) Endomorphin 1 (agonist)
SNC 80 (agonist) U-69593 (agonist) Endomorphin 2 (agonist)
DSLET (agonist) ICI-199,441 (agonist) Fentanyl citrate (agonist)
SNC121 (agonist) Nor-binaltorphimine (antagonist) (cid:12)-Funaltrexamine (antagonist)
Superfit (affinity label) DIPPA (antagonist) Naloxonazine (antagonist)
Naltrindole (antagonist) Cyprodime HBr (antagonist)
ICI-174,864 (antagonist)
BNTX (antagonist)
Naltriben (antagonist)
Table 2 Evidence for the presence of opioid receptors on primary cells of the immune systema
Cell type (cid:14) Receptor (cid:20) Receptor (cid:22) Receptor
Mouse T lymphocyte B, M – B
Mouse B lymphocyte B – B
Mouse thymocyte – – P, M
Mouse splenocyte B, M B B
Human PBLs M M –
Human T lymphocyte P, B M –
Human B lymphocyte B – –
Human granulocyte – – P, M
Human monocyte – M M
Monkey PBLs M M M
Human microglia – M –
Rat macrophage – – M
aEvidencefortheexistenceofthereceptorsisdefinedusingpharmacological(P),biochemical(B),ormolecular
biology(M)approaches(Aliceaetal.,1998;Chaoetal.,1996;Gaveriauxetal.,1995;Miller,1996;
Royetal.,1992;Wicketal.,1996)orasreviewedbyCarr(1991),Carretal.(1996).
–,Suggestseitheralackofdetectionorthattheanalysishasnotyetbeendetermined.
assays), biochemical (affinity labeling), or molecular (Roy et al., 1992) or mitogen (concanavalin A) in the
biology techniques (cloning or RT-PCR studies). case of the mouse T lymphocyte (cid:14) opioid receptor
(Miller, 1996). Furthermore, the activation of leuko-
cytes also leads to the production of endogenous
Regulation of receptor expression
opioid peptides (Blalock, 1989). Since the leukocyte-
derived opioid peptides have also been shown to be
The expression of (cid:14) and (cid:22) opioid receptors on functionally active (Blalock, 1989), there is reason to
immune cells is reportedly induced by the activation believe that autocrine regulation of receptor expres-
ofcellsbyIL-1inthecaseofthethymocyte(cid:22)receptor sion may occur as well.
Opioid (cid:22), (cid:14), and (cid:20) Receptors for Endorphins 2215
Table 3 Evidence for the presence of opioid receptors on cell lines derived from cells of the immune
systema
Cell type (cid:14) Receptor (cid:20) Receptor (cid:22) Receptor
Mouse T cell lines
EL-4 M P –
11.10 M – –
Mouse B cell line
CH27 M – –
Mouse macrophage cell line
P388d B P, B
1
Mouse R1.1 thymoma – P, M –
Human T cell lines
CEMx174 M M M
HSB2 M – –
MOLT-4 B, M M –
Human B cell line
EBV-transformed M M
Human monocyte cell line
U937 M – –
aEvidencefortheexistenceofthereceptorsisdefinedusingpharmacological(P),biochemical(B),ormolecular
biology(M)approaches(Gaveriauxetal.,1995;Chaoetal.,1996;Wicketal.,1996;Aliceaetal.,1998)or
asreviewedbyCarr(1991),Carretal.(1996).
–,Suggestseitheralackofdetectionorthattheanalysishasnotyetbeendetermined.
SIGNAL TRANSDUCTION augment T cell activation through the increase in
phosphorylation of the CD3 complex intracellular
tyrosine-activation motifs (ITAMs) and presumably
Cytoplasmic signaling cascades
the activation of the inositol trisphosphate (IP ) cas-
3
cade via ZAP-70, whereas at femtomolar levels the
Neuronal opioid receptors modify a variety of
endorphins would suppress T cell activation by
signaling cascades including cAMP through the
reducing phosphorylation.
activation of G, increases in GTPase activity, phos-
i
phatidylinositol turnover, mobilization of Ca2+, and
K+ channel activity (Childers, 1991; Chen and Yu, DOWNSTREAM GENE
1994). In a similar fashion, immune cell-derived
ACTIVATION
opioid receptors are coupled to a G protein and
i
influence K+ channel conductance and calcium
Transcription factors activated
mobilization (Carr, 1991). In addition, the endogen-
ous opioid peptide (cid:12)-endorphin has been shown to
modify CD3(cid:13) phosphorylation following phorbol The success in transfecting Jurkat T cells (which do
esterstimulation,eitherincreasingordecreasingphos- not express opioid receptors, Gaveriaux et al., 1995)
phorylation of the CD3 chain depending on the withafunctional(cid:14)opioidreceptor(Sharpetal.,1996)
concentration of the peptide (Kavelaars et al., 1990). allowed for the identification of potential transcrip-
These results suggest that the endorphins may act as tional regulatory elements involved in opioid modu-
a governor on T cell activation depending on the lation of immune function. Previous studies reported
local concentration of endogenous opioid peptide. the augmentation of IL-2 production by activated
Specifically, endorphins at mid-picomolar levels may T cells stimulated with endogenous opioid peptides
2216 Daniel J. J. Carr and J. Edwin Blalock
(Carr, 1991). In an elegant study, reporter gene con- The hypothalamic-pituitary-adrenal axis results in
structs were used to map deltorphin ((cid:14) selective the production of adrenal steroids such as gluco-
agonist)-elicited augmentation of IL-2 production by corticoids which suppress immune responses in part
(cid:14) opioid receptor-transfected Jurkat T cells to the by preventing translocation of NF(cid:20)B to the nucleus
AP-1- and NF-AT/AP-1-binding site (Hedin et al., (Baldwin, 1996). Alternatively, morphine may acti-
1997). This effect was apparently independent of vate the sympathetic/parasympathetic arm of the
calcineurin and unrelated to the elevation in [Ca2+] autonomic nervous system known to innervate
i
butrequiredpertussistoxin-sensitiveGprotein.Since lymph nodes and spleen (Felten et al., 1987) and
theNF-AT/AP-1complexisinvolvedintheinduction modify immune function through the release of
ofanumberofcytokinegenes(Rao,1994)andendo- monoamines (e.g. catecholamines) (Carr and Serou,
genous opioid peptides modify the production of a 1995).
numberofcytokines(Petersonetal., 1998),itisquite Other studies suggest that endogenous opioid pep-
possible that the NF-AT/AP-1 complex is involved. tides may supplement antimicrobial drugs or local
In addition, since leukocyte activation is primarily immune reactivity against viral infections. Studies
mediated by cytokines, it is highly probable that have suggested that met-enkephalin suppresses
opioid receptor promoters possess binding domains influenza virus infection in mice through the effects
for cytokine responsive elements. As an example, on natural killer cells and cytotoxic T lymphocytes
the (cid:22) opioid receptor promoter possesses a NF-IL6 (Burger et al., 1995). Another study has found that
domain (Min et al., 1994). met-enkephalin synergizes with azidothymidine in
blocking feline leukemia virus replication (Specter
et al., 1994). It has also been reported that endo-
Genes induced
genous opioids induce the synthesis of novel fentanyl
derivatives that possess analgesic activity in the
IL-2. absence of opioid immunosuppression (Carr and
Serou, 1995).
BIOLOGICAL CONSEQUENCES
OF ACTIVATING OR INHIBITING Phenotypes of receptor knockouts
RECEPTOR AND and receptor overexpression mice
PATHOPHYSIOLOGY
Muopioidreceptor(MOR)knockoutmicehavebeen
Unique biological effects of developed and tested for immune deviation in the
presenceandabsenceoftheclinicallyrelevant,proto-
activating the receptors
typic(cid:22)ligandmorphine.MORknockoutmiceexhibit
normal immunological endpoints including natural
The response to opioid receptor activation depends killer activity, antibody production, and mitogen-
on the location of the receptor, the type of receptor, induced lymphocyte proliferation (Gaveriaux-Ruff
and the level of activation of the cell population. etal.,1998).However,bonemarrowcellsfromMOR
Endogenous opioid peptides can either enhance or knockout mice exhibit an altered pattern of early
suppress immune function depending in part on the hematopoiesis (Tian et al., 1997). In addition, treat-
state of target cell activation and the immune param- ment with morphine had no effect on immune
eter (antibody production, natural killer activity, parameters assayed in MOR knockout mice, but sig-
cytokine synthesis) measured. Likewise, peripheral nificantly suppressed selectively measured immune
bloodmononuclearcellsfromindividualscanrespond parameters (e.g. natural killer cell activity) and
differently (sometimes completely opposite of one induced lymphoid organ atrophy in wild-type mice
another)toopioidligandsevidentinbothhumanand (Gaveriaux-Ruff et al., 1998). These results suggest
mouse populations. However, the administration of that the absence of the (cid:22) opioid receptor has no
opioid alkaloids (i.e. morphine, heroin, or fentanyl) detrimental effect on immunocompetence per se, but
tends to elicit a significant suppression of immune is directly responsible for the immunomodulatory
function primarily by opioid receptors found in the effects of exogenous morphine. Accordingly, mod-
mesencephalon (Shavit et al., 1986; Weber and Pert, ificationofimmuneresponsestoantigenormicrobial
1989). pathogens by endogenous opioid peptides does not
The activation of the ‘central’ opioid receptors necessarily involve the activation of (cid:22) opioid recep-
elicits the activation of neuroendocrine pathways. tors on immune cells.
Opioid (cid:22), (cid:14), and (cid:20) Receptors for Endorphins 2217
THERAPEUTIC UTILITY Carr, D. J. J., DeCosta, B. R., Kim, C.-H., Jacobson, A. E.,
Bost,K.L.,Rice,K.C.,andBlalock,J.E.(1990).Anti-opioid
receptor antibody recognition of a binding site on brain and
Effects of inhibitors (antibodies) leukocyteopioidreceptors.Neuroendocrinology51,552–560.
Carr,D.J.J.,Rogers,T.J.,andWeber,R.J.(1996).Therelevance
to receptors
of opioids and opioid receptors on immunocompetence and
immunehomeostasis.Proc.Soc.Exp.Biol.Med.213,248–257.
The classical pharmacological definition of the exis- Chao, C. C., Gekker, G., Hu, S., Sheng, W. S., Shark, K. B.,
Bu, D.-F., Archer, S., Bidlack, J. M., and Portoghese, P. K.
tence of opioid receptors on cells of the immune
(1996). (cid:20) opioid receptors in human microglia downregulate
system has come from the ability of opioid antag-
human immunodeficiency virus 1 expression. Proc. Natl Acad.
onists (competitive or noncompetitive) to block the Sci.USA93,8051–8056.
immunomodulatoryeffectsinastereospecificmanner Chen, Y., and Yu, L. (1994). Differential regulation by cAMP-
(Sibinga and Goldstein, 1988) (see Table 1). Anti- dependentproteinkinaseandproteinkinaseCofthe(cid:22)opioid
receptorcouplingtoaGprotein-activatedK+channel.J.Biol.
bodieshavealsobeengeneratedagainstopioidrecep-
Chem.269,7839–7842.
tors that recognize proteins expressed on cells of the
Childers,S.R.(1991).Opioidreceptor-coupledsecondmessenger
immune system. One such antibody was found to systems.LifeSci.48,1991–2003.
possess agonist activity and to recognize a putative (cid:14) Chuang, T. K., Killam Jr, K. F., Chuang, L. F., Kung, H.-F.,
classopioidreceptoronmouseleukocytes(Carretal., Sheng,W.S.,Chao,C.C.,Yu,L.,andChuang,R.Y.(1995).
Muopioidreceptorgeneexpressioninimmunecells. Biochem.
1990). A second antibody was generated against the
Biophys.Res.Commun.216,922–930.
predicted N-terminal sequence of a (cid:20) opioid receptor
Claude, P. A., Wotta, D. R., Zhang, X. H., Prather, P. L.,
and was found to act as a noncompetitive selective McGinn, T. M., Erickson, L. J., Loh, H. H., and Law, P. Y.
antagonist recognizing (cid:20) opioid receptors on U937 (1996).MutationofaconservedserineinTM4ofopioidrecep-
cells (Buchner et al., 1997). However, the use of anti- tors confers full agonistic properties to classical antagonists.
Proc.NatlAcad.Sci.USA93,5715–5719.
bodies is more apt to focus on characterizing the
Cox,B.M.,Goldstein,A.,andLi,C.H.(1976).Opioidactivityof
structural properties of the cloned receptor (e.g. apeptide,beta-lipotropin-(61-91),derivedfrombeta-lipotropin.
mapping ligand-binding domains) rather than using Proc.NatlAcad.Sci.USA73,1821–1823.
such antibodies to antagonize tolerance or chemical Evans, C. J., Keith, D., Magendzo, K., Morrison, H., and
dependence. Edwards, R. H. (1992). Cloning of a delta opioid receptor by
functionalexpression.Science258,1952–1955.
Felten, D. L., Felten, S. Y., Bellinger, D. L., Carlson, S. L.,
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