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Proc Natl Acad Sci U S A. 2001 September 25; 98(20): 11042–11046. doi: 10.1073/pnas.191352698. | PMCID: PMC58680 |
Copyright © 2001, The National Academy of Sciences ΔFosB: A sustained molecular switch for addiction Eric J. Nestler,* Michel Barrot, and David W. Self Department of Psychiatry and Center for Basic Neuroscience, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-9070 The longevity of some of the behavioral abnormalities that
characterize drug addiction has suggested that regulation of neural
gene expression may be involved in the process by which drugs of abuse
cause a state of addiction. Increasing evidence suggests that the
transcription factor ΔFosB represents one mechanism by which drugs of
abuse produce relatively stable changes in the brain that contribute to
the addiction phenotype. ΔFosB, a member of the Fos family of
transcription factors, accumulates within a subset of neurons of the
nucleus accumbens and dorsal striatum (brain regions important for
addiction) after repeated administration of many kinds of drugs of
abuse. Similar accumulation of ΔFosB occurs after compulsive running,
which suggests that ΔFosB may accumulate in response to many types of
compulsive behaviors. Importantly, ΔFosB persists in neurons for
relatively long periods of time because of its extraordinary stability.
Therefore, ΔFosB represents a molecular mechanism that could initiate
and then sustain changes in gene expression that persist long after
drug exposure ceases. Studies in inducible transgenic mice that
overexpress either ΔFosB or a dominant negative inhibitor of the
protein provide direct evidence that ΔFosB causes increased
sensitivity to the behavioral effects of drugs of abuse and, possibly,
increased drug seeking behavior. This work supports the view that
ΔFosB functions as a type of sustained “molecular switch” that
gradually converts acute drug responses into relatively stable
adaptations that contribute to the long-term neural and behavioral
plasticity that underlies addiction. Addiction research is focused
on understanding the complex ways in which drugs of abuse change the
brain to cause behavioral abnormalities that characterize addiction.
One of the critical challenges in the field is to identify relatively
stable drug-induced changes in the brain to account for those
behavioral abnormalities that are particularly long-lived. For example,
a human addict may be at increased risk for relapse even after years of
abstinence. The stability of these behavioral abnormalities has led to the
suggestion that they may be mediated, at least in part, through changes
in gene expression ( 1– 3). According to this view, repeated exposure to
a drug of abuse repeatedly perturbs transmission at particular synapses
in the brain that are sensitive to the drug. Such perturbations
eventually signal via intracellular messenger cascades to the nucleus,
where they first initiate and then maintain changes in the expression
of specific genes. A primary mechanism through which signal
transduction pathways influence gene expression is the regulation of
transcription factors, proteins that bind to regulatory regions of
genes and modify their transcription. One goal of addiction research, therefore, has been to identify
transcription factors that are altered in brain regions implicated in
addiction after chronic administration of drugs of abuse. Several such
transcription factors have been identified over the past decade ( 1– 6).
The focus of this review is on one particular transcription factor
called ΔFosB. Induction of ΔFosB by Drugs of Abuse ΔFosB, encoded by the fosB gene, is a member of the
Fos family of transcription factors, which also include c-Fos, FosB,
Fra1, and Fra2 ( 7). These Fos family proteins heterodimerize with Jun
family proteins (c-Jun, JunB, or JunD) to form active AP-1 (activator
protein-1) transcription factors that bind to AP-1 sites (consensus
sequence: TGAC/GTCA) present in the promoters of certain genes to
regulate their transcription. These Fos family proteins are induced rapidly and transiently in
specific brain regions after acute administration of many drugs of
abuse (Fig. ) ( 8– 11). Prominent regions
are the nucleus accumbens and dorsal striatum, which are important
mediators of behavioral responses to the drugs, in particular, their
rewarding and locomotor-activating effects ( 12, 13). These proteins
return to basal levels within hours of drug administration.
Very different responses are seen after chronic administration of drugs
of abuse (Fig. ). Biochemically modified isoforms of ΔFosB
(molecular mass 35–37 kDa) accumulate within the same brain regions
after repeated drug exposure, whereas all other Fos family members show
tolerance (that is, reduced induction compared with initial drug
exposures). Such accumulation of ΔFosB has been observed for cocaine,
morphine, amphetamine, alcohol, nicotine, and phencyclidine ( 11,
14– 18). There is some evidence that this induction is selective for
the dynorphin/substance P-containing subset of medium spiny neurons
located in these brain regions ( 15, 17), although more work is needed
to establish this with certainty. The 35- to 37-kDa isoforms of ΔFosB
dimerize predominantly with JunD to form an active and long-lasting
AP-1 complex within these brain regions ( 19, 20). These ΔFosB
isoforms accumulate with chronic drug exposure because of their
extraordinarily long half-lives ( 21), and therefore persist in the
neurons for at least several weeks after cessation of drug
administration. It is interesting to note that these ΔFosB isoforms
are highly stable products of an immediate early gene
( fosB). The stability of the ΔFosB isoforms
provides a novel molecular mechanism by which drug-induced changes in
gene expression can persist despite relatively long periods of drug
withdrawal. Although the nucleus accumbens plays a critical role in the rewarding
effects of drugs of abuse, it is believed to function normally by
regulating responses to natural reinforcers, such as food, drink, sex,
and social interactions ( 12, 13). As a result, there is considerable
interest in a possible role of this brain region in other compulsive
behaviors (e.g., pathological overeating, gambling, exercise, etc.).
For this reason, we examined whether ΔFosB is regulated in an animal
model of compulsive running. Indeed, the stable 35- to 37-kDa isoforms
of ΔFosB are induced selectively within the nucleus accumbens in rats
that show compulsive running behavior. †Biochemical Identity of Stable ΔFosB Isoforms As mentioned above, the ΔFosB isoforms that accumulate after
chronic administration of a drug of abuse or compulsive running show a
molecular mass of 35–37 kDa. They can be differentiated from the
33-kDa isoform of ΔFosB that is induced rapidly but transiently after
a single drug exposure (Fig. ) ( 14, 19, 22). Current evidence suggests
that the 33-kDa isoform is the native form of the protein, which is
altered to form the more stable 35- to 37-kDa products ( 19, 21).
However, the nature of the biochemical modification that converts the
unstable 33-kDa isoform into the stable 35- to 37-kDa isoforms has
remained obscure. It has been speculated that phosphorylation may be
responsible ( 11). For example, induction of ΔFosB is attenuated in
mice lacking DARPP-32, a striatal-enriched protein ( 23, 24). Because
DARPP-32 regulates the catalytic activity of protein phosphatase-1 and
protein kinase A ( 25, 26), the requirement for this protein for the
normal accumulation of the stable ΔFosB isoforms suggests a possible
role for phosphorylation in generation of these stable products. Role of ΔFosB in Behavioral Plasticity to Drugs of Abuse Insight into the role of ΔFosB in drug addiction has come
largely from the study of transgenic mice in which ΔFosB can be
induced selectively within the nucleus accumbens and other striatal
regions of adult animals ( 27, 28). Importantly, these mice overexpress
ΔFosB selectively in the dynorphin/substance P-containing medium
spiny neurons, where the drugs are believed to induce the protein. The
behavioral phenotype of the ΔFosB-overexpressing mice, which in many
ways resembles animals after chronic drug exposure, is summarized in
Table 1. The mice show augmented
locomotor responses to cocaine after acute and chronic administration
( 28). They also show enhanced sensitivity to the rewarding effects of
cocaine and morphine in place-conditioning assays ( 11, 28) and will
self-administer lower doses of cocaine than littermates that do not
overexpress ΔFosB. ‡ In
contrast, these animals show normal
conditioned locomotor sensitization to cocaine and normal spatial
learning in the Morris water maze
( 28). These data
indicate that ΔFosB increases an animal's sensitivity to cocaine and
perhaps other drugs of abuse and may represent a mechanism for
relatively prolonged sensitization to the drugs.
| Table 1Behavioral plasticity mediated by ΔFosB in nucleus
accumbens-dorsal striatum |
In addition, there is preliminary evidence that the effects of ΔFosB
may extend well beyond a regulation of drug sensitivity per
se to more complex behaviors related to the addiction process.
Mice expressing ΔFosB work harder to self-administer cocaine in
progressive ratio self-administration assays, suggesting that ΔFosB
may sensitize animals to the incentive motivational properties of
cocaine and thereby lead to a propensity for relapse after drug
withdrawal. ‡ ΔFosB-expressing mice also show
enhanced anxiolytic effects of alcohol, § a
phenotype that has been associated with increased alcohol intake in
humans. Together, these early findings suggest that ΔFosB, in
addition to increasing sensitivity to drugs of abuse, produces
qualitative changes in behavior that promote drug-seeking behavior.
Thus, ΔFosB may function as a sustained “molecular switch” that
helps initiate and then maintain crucial aspects of the addicted state.
An important question under current investigation is whether ΔFosB
accumulation during drug exposure promotes drug-seeking behavior after
extended withdrawal periods, even after ΔFosB levels have normalized
(see below). Adult mice that overexpress ΔFosB selectively within the nucleus
accumbens and dorsal striatum also exhibit greater compulsive running
compared with control littermates. † These
observations raise the interesting possibility that ΔFosB
accumulation within these neurons serves a more general role in the
formation and maintenance of habit memories and compulsive behaviors,
perhaps by reinforcing the efficacy of neural circuits in which those
neurons function. ΔFosB accumulates in certain brain regions outside the nucleus
accumbens and dorsal striatum after chronic exposure to cocaine.
Prominent among these regions are the amygdala and medial prefrontal
cortex ( 15). A major goal of current research is to understand the
contributions of ΔFosB induction in these regions to the addiction
phenotype. Earlier work on fosB knockout mice revealed that these
animals fail to develop sensitization to the locomotor effects of
cocaine, which is consistent with the findings of the
ΔFosB-overexpressing mice mentioned above ( 22). However, the
fosB mutants showed enhanced sensitivity to cocaine's acute
effects, which is inconsistent with these other findings.
Interpretation of findings with the fosB mutants, though, is
complicated by the fact that these animals lack not only ΔFosB, but
full-length FosB as well. Moreover, the mutants lack both proteins
throughout the brain and from the earliest stages of development.
Indeed, more recent work supports conclusions from the ΔFosB
overexpressing mice: inducible overexpression of a truncated mutant of
c-Jun, which acts as a dominant negative antagonist of ΔFosB,
selectively in nucleus accumbens and dorsal striatum shows reduced
sensitivity to the rewarding effects of cocaine. ¶
These findings emphasize the caution that must be used in interpreting
results from mice with constitutive mutations and illustrate the
importance of mice with inducible and cell type-specific mutations in
studies of plasticity in the adult brain. Because ΔFosB is a transcription factor, presumably the protein
causes behavioral plasticity through alterations in the expression of
other genes. ΔFosB is generated by alternative splicing of the
fosB gene and lacks a portion of the C-terminal
transactivation domain present in full-length FosB. As a result, it was
originally proposed that ΔFosB functions as a transcriptional
repressor ( 29). However, work in cell culture has demonstrated clearly
that ΔFosB can either induce or repress AP-1-mediated transcription
depending on the particular AP-1 site used ( 21, 29– 31). Full-length
FosB exerts the same effects as ΔFosB on certain promoter fragments,
but different effects on others. Further work is needed to understand
the mechanisms underlying these varied actions of ΔFosB and FosB. Our group has used two approaches to identify target genes for ΔFosB.
One is the candidate gene approach. We initially considered
α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate
receptors as putative targets, given the important role of
glutamatergic transmission in the nucleus accumbens. Work to date has
indicated that one particular AMPA glutamate receptor subunit, GluR2,
may be a bona fide target for ΔFosB (Fig.
). GluR2 expression, but not the
expression of other AMPA receptor subunits, is increased in nucleus
accumbens (but not dorsal striatum) upon overexpression of ΔFosB
( 28), and expression of a dominant negative mutant attenuates the
ability of cocaine to induce the protein. ¶ In
addition, the promoter of the GluR2 gene contains a consensus AP-1 site
that binds ΔFosB ( 28). Overexpression of GluR2 in the nucleus
accumbens, by use of viral-mediated gene transfer, increases an
animal's sensitivity to the rewarding effects of cocaine, thereby
mimicking part of the phenotype seen in the ΔFosB-expressing mice
( 28). Induction of GluR2 could account for the reduced
electrophysiological sensitivity of nucleus accumbens neurons to AMPA
receptor agonists after chronic cocaine administration ( 32), because
AMPA receptors containing GluR2 show reduced overall conductance and
reduced Ca 2+ permeability. Reduced responsiveness
of these neurons to excitatory inputs may then enhance responses to a
drug of abuse. However, the ways in which dopaminergic and
glutamatergic signals in nucleus accumbens regulate addictive behavior
remain unknown; this will require a neural circuit level of
understanding, which is not yet available.
Another putative target for ΔFosB is the gene encoding dynorphin. As
stated earlier, dynorphin is expressed in the subset of nucleus
accumbens medium spiny neurons that show induction of ΔFosB.
Dynorphin appears to function in an intercellular feedback loop: its
release inhibits the dopaminergic neurons that innervate the medium
spiny neurons, via κ opioid receptors present on dopaminergic nerve
terminals in the nucleus accumbens and also on cell bodies and
dendrites in the ventral tegmental area (Fig.
) ( 33– 35). This idea is consistent with
the ability of a κ receptor agonist, upon administration into either
of these two brain regions, to decrease drug reward ( 35). Recent work
has indicated that ΔFosB decreases the expression of
dynorphin, ‖ which
could contribute to the enhancement of reward mechanisms seen with
ΔFosB induction. Interestingly, another drug-regulated transcription
factor, CREB (cAMP response element binding protein) ( 2, 3), exerts the
opposite effect: it induces dynorphin expression in the nucleus
accumbens and reduces the rewarding properties of cocaine and morphine
( 4). ** Because drug-induced activation of
CREB dissipates rapidly after drug administration, such reciprocal
regulation of dynorphin by CREB and ΔFosB could explain the
reciprocal behavioral changes that occur during early and late phases
of withdrawal, with negative emotional symptoms and reduced drug
sensitivity predominating during early phases of withdrawal, and
sensitization to the rewarding and incentive motivational effects of
drugs predominating at later time points.
The second approach used to identify target genes for ΔFosB involves
DNA microarray analysis. Inducible overexpression of ΔFosB increases
or decreases the expression of numerous genes in the nucleus accumbens
( 36). Although considerable work is now needed to validate each of
these genes as physiologic targets of ΔFosB and to understand their
contribution to the addiction phenotype, one important target appears
to be Cdk5 (cyclin-dependent kinase-5). Thus, Cdk5 was initially
identified as ΔFosB-regulated by use of microarrays, and later shown
to be induced in nucleus accumbens and dorsal striatum after chronic
cocaine administration ( 37). ΔFosB activates the cdk5 gene
via an AP-1 site present within the gene's promoter ( 36). Together,
these data support a scheme wherein cocaine induces Cdk5 expression in
these brain regions via ΔFosB. Induction of Cdk5 appears to alter
dopaminergic signaling at least in part via increased phosphorylation
of DARPP-32 ( 37), which is converted from an inhibitor of protein
phosphatase-1 to an inhibitor of protein kinase A upon its
phosphorylation by Cdk5 ( 26). Role of ΔFosB in Mediating “Permanent” Plasticity to Drugs
of Abuse Although the ΔFosB signal is relatively long-lived, it is not
permanent. ΔFosB degrades gradually and can no longer be detected in
brain after 1–2 months of drug withdrawal, even though certain
behavioral abnormalities persist for much longer periods of time.
Therefore, ΔFosB per se would not appear to be able to
mediate these semipermanent behavioral abnormalities. The difficulty in
finding the molecular adaptations that underlie the extremely stable
behavioral changes associated with addiction is analogous to the
challenges faced in the learning and memory field. Although there are
elegant cellular and molecular models of learning and memory, it has
not to date been possible to identify molecular and cellular
adaptations that are sufficiently long-lived to account for highly
stable behavioral memories. Indeed, ΔFosB is the longest-lived
adaptation known to occur in adult brain, not only in response to drugs
of abuse, but to any other perturbation (that doesn't involve lesions)
as well. Two proposals have evolved, both in the addiction and learning
and memory fields, to account for this discrepancy. One possibility is that more transient changes in gene
expression, such as those mediated via ΔFosB or other transcription
factors (e.g., CREB), may mediate more long-lived changes in neuronal
morphology and synaptic structure. For example, an increase in the
density of dendritic spines (particularly an increase in two-headed
spines) accompanies the increased efficacy of glutamatergic synapses at
hippocampal pyramidal neurons during long-term potentiation ( 38– 40),
and parallels the enhanced behavioral sensitivity to cocaine mediated
at the level of medium spiny neurons of the nucleus accumbens ( 41). It
is not known whether such structural changes are sufficiently
long-lived to account for highly stable changes in behavior, although
the latter persist for at least 1 month of drug withdrawal. Recent
evidence raises the possibility that ΔFosB, and its induction of
Cdk5, is one mediator of drug-induced changes in synaptic structure in
the nucleus accumbens (Fig.
). ‡‡ Thus, infusion of a
Cdk5 inhibitor into the nucleus accumbens
prevents the ability of repeated cocaine
exposure to increase dendritic spine density in this region. This is
consistent with the view that Cdk5, which is enriched in brain,
regulates neural structure and growth (see refs. 36 and 37). It is
possible, although by no means proven, that such changes in neuronal
morphology may outlast the ΔFosB signal itself.
Another possibility is that the transient induction of a transcription
factor (e.g., ΔFosB, CREB) leads to more permanent changes in gene
expression through the modification of chromatin. These and many other
transcription factors are believed to activate or repress the
transcription of a target gene by promoting the acetylation or
deacetylation, respectively, of histones in the vicinity of the gene
( 42). Although such acetylation and deacetylation of histones can
apparently occur very rapidly, it is possible that ΔFosB or CREB
might produce longer-lasting adaptations in the enzymatic machinery
that controls histone acetylation. ΔFosB or CREB may also promote
longer-lived changes in gene expression by regulating other
modifications of chromatin (e.g., DNA or histone methylation) that have
been implicated in the permanent changes in gene transcription that
occur during development (see refs. 42 and 43). Although these
possibilities remain speculative, they could provide a mechanism by
which transient adaptations to a drug of abuse (or some other
perturbation) lead to essentially life-long behavioral consequences. This work was supported by grants from the National Institute on
Drug Abuse. | | AP-1 | activator protein-1 | | | AMPA | α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid | | | CREB | cAMP
response element binding protein | | | Cdk5 | cyclin-dependent kinase-5 |
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