Cytokine purine interactions in behavioral depression in rats

1. Cytokine-Purine Interactions in
Behavioral Depression in Rats
THOMAS R. MINOR 1, QINGJUN HUANG2, AND ELIZABETH A. FOLEY1
lDepartment of Psychology, University of California, Los Angeles, Los Angeles CA 90095-1563
2Psychoneuroimmunology Laboratory, Bethune Military Medical College, 450 West Zhongshan
Road, Shijiazhuang, Hebei 050081, China
This paper reviews recent findings from our laboratories concerning metabolic and immune mediators of behavioral depression in rats. Specifically, a single injection of 6 mg/kg
of reserpine substantially increases behavioral depression, as evidenced by an increase in
the amount of time spent floating by independent groups of rats tested for swim performance at various times during the next week. The behavioral impairment consists of two
components. An early component emerges one hour after reserpine treatment and persists
for about 24 hours. The deficit is not reversed by intracranial ventricular infusion of the
receptor antagonist for interleukin-l{I (IL-I~). A second, late-component deficit appears
approximately 48 hours after reserpine treatment and recovers within a week. Late-component
depression is reversed by central infusion of the IL-16 receptor antagonist, and is mimicked by central infusion of the proinflanunatory cytokine. Importantly, both early and late
components of reserpine-induced depression and IL-1 ~ induced depression are reversed by
a systemic injection of the highly selective A2A adenosine receptor antagonist 8-(3Chlorostyryl) caffeine. These data are discussed in terms of the overlap in the conservationwithdrawal reaction during sickness, traumatic stress, and major depression and the regional
contribution of purines and cytokines to the organization of this reaction in the brain.
RESERPINEIS ANalkaloid extract from the root of Rauwolfia serpentina, a climbing shrub
indigenous to India. The compound had a long history of use in the Orient as a tranquilizing agent before being introduced in the United States in the early 1950s as a treatment for
hypertension (Cooper, Bloom & Roth, 1978; Gerber & Nies, 1990; Rech & Moore, 1971).
The extract reduces both cardiac output and peripheral vascular resistance by depleting
stores of biogenic amines in the central and autonomic nervous systems. Specifically,
reserpine binds irreversibly to storage vesicles in monoaminergic neurons (Norn & Shore,
1999; Schwartz, 1981). The vesicles become "leaky," resulting in seepage of transmitter
into the cytoplasm, where it is either destroyed by intraneuronal monoamine oxidase or
diffuses into the synaptic cleft. The end result is that little or no active transmitter is
released at the synapse following depolarization. Recovery from the effects of reserpine
requires synthesis of new storage vesicles, which can take several days to accomplish after
discontinuing treatment (Cooper, Konkol, & Breese, 1978; Ponzio, Achilli, Calderini &
Ferretti, 1984).
Address for Correspondence: Thomas R. Minor, Ph.D., Department of Psychology, University of California, Los
Angeles, Los Angeles, CA 90095-1563, E-mail: Minor@psych.ucla.edu.
Integrative Physiological & Behavioral Science, July-September 2003, Vol. 38, No. 3, 189-202.
189

2. 190
MINOR, HUANG, AND FOLEY
The historic importance of reserpine is more related to its unwanted side effects than to
its efficacy as an antihypertensive or tranquilizing agent. Unfortunately, a significant portion of the population undergoing reserpine treatment for hypertension developed symptoms of major depression. These symptoms were severe enough to require antidepressant
drug treatment and, at times, hospitalization. This observation, along with findings a few
years later that monoamine oxidase inhibitors and tricyclic antidepressants enhance brain
biogenic amines, severed as the empirical cornerstone of catecholamine (Schildkraut,
1965), and later, monoamine theories of depression (Akiskal & McKinney, 1975; Bunney
& Davis, 1965).
This paper reviews recent research from our laboratories (Haung & Minor, 2003 a, b, c;
Jiang, Minor, & Huang, 2003). We revisited the animal literature on reserpine to ask
whether it is the depletion of brain monoamines per se, or some downstream consequence
of reserpine treatment, that precipitates a depressive episode. Despite the promise of the
original catecholamine theory, empirical evidence linking any of the biogenic amines and
major depression is less than convincing. As an alternative, we focused on the potential
contribution of two brain-signaling pathways, one mediated by the purine nucleoside adenosine and the other involving the proinflammatory cytokine interleukin-1 ~ (IL-1 ~). Both
of these pathways are capable of producing a conservation-withdrawal reaction (Engle &
Schmale, 1971) and both are plausibly engaged by reserpine treatment. This reaction
corresponds to the fatigue component of major depression (Field & Reite, 1984; Minor et
al., 1994; Woodson et al., 1998). A conservation-withdrawal reaction unconditionally follows periods of intense catabolic output (traumatic psychological stress, injury, or severe
illness). The sensory unresponsiveness, cognitive dullness, and behavioral depression that
characterize this state are assumed to be adaptive responses that husband limited resources
and facilitate the recovery of metabolic homeostasis.
Brain Adenosine Signaling.
Adenosine signaling links cellular excitability to energy state and is actively engaged by
challenges to metabolic homeostasis (Van Wylen et al., 1986; Phillis et al., 1987). The
nucleoside exerts very potent inhibition on excitatory transmission in the brain as a compensatory reaction to neural energy failure (Fredholm & Dunwiddie, 1988; Newby, 1984;
Meghji, 1991; Stone, 1981). Adenosine is extruded into extracellular space, or hydrolyzed
from extracellular nucleotides, whenever the rate of adenosine triphosphate (ATP) hydrolysis exceeds the synthesis rate (Meghji, 1991; Newby, 1984; White & Hoehn, 1991).
Such an imbalance of the energy supply/demand ratio can result from excessive neural
activation or from a shortage in brain glucose or oxygen. The extracellular nucleoside
binds to specific adenosine receptors (A1, A2A, A2B, & A3), which are widely distributed
on pre- and post-synaptic membranes and in the brain microvascular bed (Daly, 1990;
Linden, 1991; Rail, 1990). Adenosine interacts with a number of cellular effector systems
via these receptors to decrease membrane excitability and inhibit transmitter release,
thereby decreasing metabolic demand in the target neuron (Burns, 1991). Adenosine also
acts at the system level to produce a number of changes that protect neural tissue from the
potentially excitotoxic effects of activation in the absence of sufficient energy (Dragunow,
1988; Harms, Wardeh, 1979; Marangos, 1991; Milusheva et al., 1990; Novelli, Reilly,
Lyskoi, & Henneberry, 1988; Daval & Nicolas, 1998).
Adenosine signaling plays a crucial role in mediating the transition from an anxious/
agitated state to one of behavioral depression in the learned helplessness paradigm (Minor,

3. CYTOK1NE-PURINE INTERACTIONS IN BEHAVIORAL DEPRESSION
191
Chang, & Winselow, 1994; Minor, Winselow & Chang, 1994; Minor & Hunter 2002) and
in mediating immobility in a forced swim test in mice (El Yacoubi et al., 2001). Adenosine
also is plausibly activated by reserpine treatment. One of the conditions under which
adenosine exerts potent compensatory inhibition is during excessive neural activation.
Large amounts of transmitter are likely to be released upon initial reserpine treatment and
during the process of depleting the biogenic amines. The resulting excitation might be
sufficient to compromise metabolic homeostasis and provoke adenosine-mediated inhibition as a compensatory mechanism. This possibility is supported by the similarity in the
conservation-withdrawal responses following reserpine treatment and inescapable shock.
Cytokine Signaling
Systemic administration of lipopolysaccharide (LPS), the active fragment of endotoxin
from gram-negative bacteria, induces the synthesis of proinflammatory cytokines in peripheral macrophages---interleukin-ll3 (IL-I[3), IL-6, and tumor necrosis factor (TNFct)
(Roitt, 1998). Kuppfer cells in the liver also express IL-I~ as a consequence of LPS
administration, and may serve as the primary immune-to-brain communication pathway.
This signal is transferred via the vagal nerve complex to the brain nucleus tractus solitarius
(NTS) where IL-113 is then expressed. The cytokine also is expressed relatively quickly
thereafter in a variety of other brain nuclei, particularly in the hypothalamus (Larson &
Dunn 200; Fleshner, Goehler, Hermann, Relton, Maier, & Watkins, 1995; Wong et al.,
1997).
IL-l[3 binds to specific receptors distributed throughout the brain to induce sickness
behavior: lethargy, hypoactivity, decreased libido, anorexia, anhedonia, and increased
sleep (Dantzer, 2001; Larson, & Dunn, 2001). This dramatic shift in ongoing activity,
along with the induction of fever, is assumed to be a highly adaptive strategy to fight infection.
Symptoms of sickness behavior and major depression overlap considerably (Maes,
1995; Pollack & Yirmiya, 2002; Yirmiya, 1996; Zacharko et al., 1997). In this regard,
systemic administration of endotoxin not only increases brain concentrations of IL- 1[I, but
also produces swim deficits (del Ceerro & Borrell, 1990) and other experimental indices of
depression (Yirmiya, 1996). These LPS-induced ailments are reversed by chronic (but not
acute) treatment with tricyclic antidepressants (Yirmiya, 1996). More important for the
present purposes is the recent finding that LPS-induced swim deficits are reversed by
systemic administration of an AaA receptor antagonist (El Yacoubi et al., 2002). These data
suggest that there is an important interaction between purine (adenosine) and cytokine (ILll3) signaling in one model of behavioral depression.
Time Course for Reserpine-Induced Depression
One of the historic problems in creating a convincing case for the biogenic amines in
reserpine-induced depression is that the time course for monoamine depletion does not fit
the time course for behavioral impairment. For instance, Bean et al. (1989) reported that
dopamine (DA) depletion in striatum, nucleus accumbens, and frontal cortex occurred
rapidly and reached a maximum about six hours after an ip injection of 5 mg/kg of
reserpine. DA levels remained at floor levels for about 18 hours after the injection and then
increased thereafter, with complete recovery occurring within 48 hours. Behavioral depression typically lasts considerably longer, although the recovery time course varies to some
degree with the specifics of the test measure.

4. 192
MINOR, HUANG, AND FOLEY
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F]6. 1. Recovery time course for reserpine-induced depression. Rats were injected with DMSO vehicle or 6
mg/kg of reserpine and then tested for swim performance 1, 24, 48, 72, or 168 hours later.
Our first objective in this line of work was to determine the specific recovery time
course in our measure of behavioral depression following reserpine treatment. We injected
groups of rats (n = 10 per group) with a single 6 mg/kg, intraperitoneal (ip) dose of
reserpine or DMSO vehicle. We used a single large dose because prior research had shown
that gradual depletion of the biogenic amines with repeated small reserpine doses does not
produce behavioral depression in rats. This effect only occurs with the rapid depletion
associated with a large dose (Cooper et al., 1978). The dose used in the time course
experiment was determined empirically in work not shown here.
Rats were tested in a swim task one, 24, 48, 72, or 168 hours after drug treatment. The
swim task consisted of placing a rat in a container of room-temperature water for 15
minutes. Behavior was videotaped and later scored for the amount of time spent floating
(complete absence of movement) or swimming. We equipped each rat with a set of Huang
water wings, a piece of Styrofoam attached to the rat's back with Velcro. Although floating
might be seen as an adaptive response in this task, we found early on that reserpine-treated
rats sank to the bottom of the container and would have drowned if not for the water wings.
Figure 1 shows mean floating time in each of the 10 groups. Vehicle controls (striped
bars) consistently spent about half of the test session floating, regardless of the time of
testing. Reserpine (solid bars) resulted in a large deficit in swim performance, with close to
a two-fold increase in floating times, relative to controls. Impairment was evident as early
as one hour after drug treatment and persisted for at least 72 hours. The deficit recovered
within a week.
Early and Late Components of the Deficit
Although reserpine produced deficits that were equal in magnitude across the first 72
hours of testing (see Figure 1), recent evidence suggests that different mediators might
emerge at different times to impair performance. For instance, over-activation of glutamate
neurons resulting from direct infusion of the excitatory amino acid transmitter into frontal

5. C Y T O K I N E - P U R I N E I N T E R A C T I O N S IN B E H A V I O R A L D E P R E S S I O N
193
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FIc. 2. Effects of the interleukin-lbeta receptor antagonist (IL-113ra) on swim performance 1 hour after
reserpine treatment. Rats initially were injected with DMSO (2 groups) or 6 mg/kg of reserpine (3 groups).
Forty-five minutes later one DMSO group received icv infusion of the IL-11~raand served as a control for
untoward effect of the antagonistduring later swim testing. Additionally,one reserpinetreated group received
icy infusion saline vehicle (Res+sVeh) and one reserpine group received icv infusion of 6 txg of the IL-l~lra
(Res+IL-113ra).Swim testing occurredin all groups 15 minuteslater.
cortex produces helplessness behavior (Hunter, Balleine, & Minor, 2003; Petty,
McChesney, & Kramer, 1985). Whereas behavioral impairment occurring one hour after
the injection in this paradigm is mediated by adenosine, behavioral depression at longer
post-injection times (e.g., 48 or 72 hours) is not (Hunter at al., 2003). These data suggested
that it would be prudent to test potential mediators of reserpine-induced depression at
different time points. We chose one hour and 48 hours, respectively, as representatives of
the early and late components of the recovery time course.
We assessed the potential contribution of the pro-inflammatory cytokine IL-115 to reserpine-induced depression in two experiments. Given that a peripheral injection of endotoxin
produces an IL-1 B-mediated deficit in swim performance, and that behavioral depression is
a component of sickness behavior, the proinflammatory cytokine seems to be a plausible
mediator of reserpine-induced swim deficits. If so, then it should be possible to reverse the
behavioral malaise produced by reserpine by administering the naturally occurring receptor
antagonist for IL-11~ directly into the brain.
We tested this possibility by stereotaxically implanting five groups of 10 rats each with
guide cannulae in the right lateral ventricle. After recovering from surgery, three groups
were injected with reserpine and two groups were injected with DMSO vehicle. Forty-five
minutes later, one of the DMSO-treated groups received a 6 Ixg intracranial ventricular
(icv) infusion of the IL-113 receptor antagonist (Group IL-l~ra) and served as a control for
any untoward effects of the receptor antagonist on swim performance during later testing.
One reserpine-treated group also received the IL-l~ra (Res+IL-1Dra) and another received
saline vehicle (Res+sVeh). The other reserpine (Res) and DMSO groups received sham
injections at this time. All groups were tested for swim performance 15 minutes later.
Figure 2 shows mean floating time in each of the five groups. Reserpine-treated rats
(Group Res) showed a large swim deficit relative to the DMSO control one hour after the

6. 194
MINOR, HUANG, AND FOLEY
15
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FI6. 3. Effects of the IL-ll3ra on swim performance 48 hours after reserpine treatment. Rats initially were
injected with DMSO (1 group) or 6 mg/kg reserpine (3 groups), and then tested for swim performance 48 hours
later. Fifteen minutes before swim testing one reserpine treated group received icy infusion of saline vehicle
(Res+sVeh) and one reserpine group received icy infusion of the IL-1 ~ra (Res+IL-1 ~ra). Groups DMSO and
Res received sham icv injections at this time.
injection. Icv injection of the IL-113ra had no untoward effect on swim performance (Group
IL-1 ~ra). The antagonist also failed to improve swim performance in reserpine-treated rats
(Group Res+sVeh vs Res+IL-113ra). Although we have not conducted a complete experiment, we have tested enough rats to be reasonably certain that the IL-l~ra also fails to
improve performance 24 hours after reserpine treatment. Thus, IL-I~ does not contribute
to the early component of the reserpine-induced deficit.
Next, we assessed the contribution of IL-113 to the behavioral depression occurring 48
hours after reserpine treatment. We randomly assigned rats to one of four groups of 10 rats
each and implanted them with a guide cannula in the right lateral ventricle. One week after
surgery, one group received an ip injection of DMSO vehicle and three groups received an
ip injection of 6 mg/kg reserpine. All rats were tested for swim performance 48 hours later.
Fifteen minutes before testing, one reserpine group received icv infusion of saline vehicle
(Res+sVeh) and one reserpine group received icv infusion of 6 ~tg of the IL-113ra (Res+ILll3ra). The other two groups received sham injections (Groups DMSO & Res).
Figure 3 shows mean floating time during the swim test in each of the four groups. An
injection of reserpine produced a large increase in floating time 48 hours later (Groups Res
vs DMSO). Icv infusion of saline 15 minutes before testing produced a slight (but not
statistically significant) increase in floating time (Group Res+sVeh). Most important, infusion of the IL-1 l]ra 15 minutes before testing substantially improved test performance such
that there was no statistical difference between Group Res+IL-1l~ra and Group DMSO.
Although we have not completed an entire experiment at the 72-hour point, we have tested
enough rats to be confident that IL-I[~ also contributes to swim deficits 72 hours after
reserpine treatment.
These experiments provide clear evidence for both an early and late component to
reserpine-induced depression, The early component is not reversed by the IL-1 [3ra, and is
not mediated by the proinflammatory cytokine. Indeed, one hour is too brief a period of
time after reserpine treatment for the immune system to be activated and an immune-tobrain signal to be generated (Brandwein, 1986; Dantzer, 2001; Goehler et al., 2000). By

7. CYTOKINE-PURINE INTERACTIONS IN BEHAVIORAL DEPRESSION
195
contrast, the late component of the deficit is reversed by central blockade of IL-1 ~ receptors, and therefore, clearly depends upon the induction of IL- 1~ in brain.
Adenosine Signaling
Although the long-term consequence of reserpine treatment is an inability to release
monoamine transmitters upon neuronal depolarization, the initial effect of the drug is
likely to be excessive and unregulated transmitter release, resulting in neuronal overexcitation. These are precisely the conditions under which adenosine is recruited as a
homeostatic and neuroprotective mechanism (Dragunow, 1988; Marangos, 1991; Meghji,
1991; Phillis et al., 1987). As noted earlier, adenosine is a well-established mediator of
behavioral depression in the learned helplessness paradigm (Minor et al., 1994 a, b).
We assessed the potential contribution of adenosine signaling to the early and late
components of reserpine depression in a lengthy series of experiments. Rats were treated
with DMSO or reserpine and then tested for swim performance one or 48 hours later.
Attempts to reverse impairment by treating rats with the selective A I antagonist 8Phenyltheophylline (8-PT), the A2s antagonist alloxazine (AX), or the A 3 antagonist
MRS-1220 15 minutes before swim testing were unsuccessful. However, the nonselective
antagonist caffeine, and the moderately selective A2 antagonist 3,7-Dimethyl-1propargylxanthine (DMPX), reversed both early and late deficits in a dose-dependent
manner. Below we review evidence that swim deficits following reserpine treatment are
mediated specifically at A2A receptors, regardless of the time of testing.
Rats were assigned randomly to one of six groups of 10 rats each. We injected four
groups with 6 mg/kg reserpine. Two other groups were injected with DMSO vehicle.
Forty-five minutes later, one DMSO-treated group was injected with 1.0 mg/kg of the
selective A2A antagonist 8-(3-Chlorostyryl) caffeine (CSC) and served as a control during
later swim testing for any untoward effects of the drug. Reserpine-treated groups received
an ip injection of 0, 0.01, 0.1, or 1.0 mg/kg of CSC. All rats were tested for swim
performance 15 minutes later.
Figure 4 shows mean floating time in each of the six groups. CSC alone had no
untoward effects on swim performance relative to the DMSO control. Pretreatment with
reserpine one hour before testing produced a large swim deficit, an effect that was completely reversed by CSC in a dose-dependent manner. Thus the early, IL-1 E-independent
deficit is mediated at A2A receptors and seems to parallel most closely the time course for
depletion of brain monoamines (Bean et al., 1989).
We assessed the effects of CSC on swim performance 48 hours after reserpine treatment
to determine whether the late component of the deficit also depends on activation of
adenosine A2A receptors. Rats were assigned randomly to one of three groups of 10 rats
each. One group was injected with DMSO vehicle and two groups were injected with 6
mg/kg reserpine. Swim testing occurred 48 hours later. Fifteen minutes before testing, one
reserpine-treated group was injected with DMSO vehicle, and one reserpine group was
injected with 1.0 mg/kg of CSC, the dose that was most effective in reversing the early
deficit in the previous experiment.
Figure 5 shows mean floating times in each of the three groups. As in previous work,
reserpine produced a large increase in floating time 48 hours later. The deficit was completely reversed by pretreatment with 1.0 mg&g of the CSC. These data clearly indicate
that the late component of reserpine-induced depression also depends on A2A receptor
activation. The data also obviate an important cytokine-purine interaction.

8. 196
MINOR, HUANG,AND FOLEY
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Fro. 4. Effects of the selective A2A antagonist 8-(3-Chlorostyryl)caffeine(CSC) on swim performance 1
hour after reserpine treatment. Rats were injected with DMSO (1 group), reserpine (4 groups), or nothing (1
group). Forty-five minutes later the untreated group received an ip injection of lmg/kg CSC and served as a
control for any untoward effects of the drug on swim performance. Additionally reserpine treated groups
received an ip injection of 0, 0.01, 0.1, or 1.0 mg/kg of CSC and swim tested 15 minutes later.
IL- l fl Induced Behavioral Depression
We also have conducted experiments that provide additional evidence for a role of IL1~1 in reserpine depression and explore more fully the cytokine-purine interaction. The
foregoing experiments suggest that treatment with reserpine produces some condition (e.g.,
cell damage in the periphery or brain) that increases the expression of IL-1 ~l in the brain.
Adenosine was recruited, perhaps as a compensatory feedback on the proinflammatory
cytokine (Bshesh et al., 2002; Trincavelli, Costa, Tuscano, Lucacchini, & Martin, 2002),
and activates extracellular A2A receptors to produce a swim deficit. Although much detail
is missing in this hypothetical chain of events, several outcomes should be obtained if the
basic sequence is correct. For instance, it should be possible to produce an immediate
swim deficit by directly infusing IL-11~ into the brain.
To test this possibility, we randomly assigned rats to one of five groups of 10 rats each.
Each rat initially was implanted with a guide cannula in the right lateral ventricle. One
week after surgery, one group received icy infusion of saline vehicle (3 B1) and three
groups received a 2 ng infusion of IL-I~. Swim testing occurred one hour later. Ten
minutes before testing, the as-yet untreated group received a 6 Bg infusion of the IL-113ra
and served as a control for any untoward effects of the antagonist during later swim
testing. One IL-1 ~ treated group received an infusion of saline vehicle and another IL-113
group received icv infusion of 6 Bg of the IL-1 ~ra. The other two groups received sham
icv injections at this time.
Figure 6 shows mean floating time during the swim test in each of the five groups. ICV
infusion of IL-1 ~ produced a large swim deficit one hour later, as evidenced in the difference between Groups sVeh and IL-I~. Central infusion of the IL-l~ra 15 minutes before
testing had no untoward effect on swim performance. More important, infusion of the

9. CYTOKINE-PURINE INTERACTIONS IN BEHAVIORAL DEPRESSION
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with DMSO (1 group) or reserpine (2 groups), and then tested for swim performance 48 hours later. Fifteen
minutes before swim testing one reserpine group received an injection of DMSO (Res+DMSO), and the other
reserpine group received and injection of 1.0 mg/kg 8-(3-Chlorostyryl)caffeine (Group Res+CSC).
receptor antagonist in IL-l-treated rats completely eliminated any evidence of a swim
deficit, indicating that the cytokine is capable of producing a swim deficit in the near term.
We also have assessed whether the IL-113 deficit depends on an interaction with A2A
adenosine receptors. Rats were assigned randomly to one of six groups of 10 rats each. All
rats underwent stereotaxic surgery to implant a guide cannula in the right lateral ventricle.
After one week of recovery, one group received icv infusion of saline vehicle (Group
sVeh) and five groups received icv infusion of 2 ng of IL- 1~. Forty-five minutes later, one
IL-113 group received an ip injection of DMSO (Group IL-I~+DMSO), and another IL-113
group received a 1.0 mg/kg ip injection of the A2A receptor antagonist CSC (Group ILI~I+CSC). Because we did not have a complete adenosine receptor profile for the IL-I~
swim deficit, we also assessed the effects of the nonselective adenosine receptor antagonist
caffeine. If CSC were ineffective in reversing the IL-113 deficit, the comparison to caffeine
would give us an indication of whether some other adenosine receptor might be involved
in the cytokine deficit. Thus, one IL-1 ~l group received an ip injection of caffeine vehicle
(10% alcohol, 40% propylene glycol, 50% H20; Group IL-1 ~l+cVeh), and one IL-113 group
received an ip injection of 7 mg/kg of caffeine (Group IL-1 ~+Caf). Groups sVeh and ILl~l received sham ip injections at this time. Swim testing occurred 15 minutes later.
Figure 7 shows mean floating time in the swim test in each of the six groups. Infusion of
IL-I~ produced a large increase in floating time one hour later (Groups sVeh versus IL1~). The magnitude of the deficit was not altered by ip injection of the caffeine vehicle
(Group IL-113+cVeh) or DMSO (Group IL-113+DMSO) vehicle 15 minutes before testing.
Importantly, the cytokine deficit was completely eliminated by pretest treatment with the
nonselective adenosine receptor antagonist caffeine (Group IL-l~+Caf) and the highly
selective A2A antagonist CSC (Group IL-I~+CSC). Thus, the IL-I~ deficit, as with the
other deficits described above, requires activation of an adenosine A2A receptor.

10. 198
MINOR, HUANG, AND FOLEY
I-HOUR POST IL-l~6 TREATMENT
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Flc. 6. Effects of the IL- 1lira on swim performance 1 hour after icy infusion of 1L-113. One group received
an icv infusion of saline vehicle (Group sVeh), and 3 groups received an icy infusion of 2 ng of IL-1I~, and one
group received nothing. Forty-five minutes later, the as of yet untreated group received a 6 ktg infusion of the
IL-l~ra and served as a control for any untoward effect of the drug on later swim performance. Additionally,
one IL-1I] group received icv infusion of saliene vehicle (Group IL-l~+sVeh), while another IL-1I] group
received an icv infusion of the 1L-libra (Group IL-l_+ra). Groups sVeh and IL-11] received sham treatment at
this time. Swim testing occured 15 minutes later.
Overview
Even though reserpine's ability to induce depressive symptomology in human and animal populations provided the empirical foundation of catecholamine/monoamine theories
of depression (Akiskal & McKinney, 1975; Bunney & Davis, 1965; Schildkraut, 1965),
the fit between data and theory was never particularly close. The main problem is that the
time course for behavioral malaise generally does not match the time course for biogenic
amine depletion.
The present data provide a clear answer as to why this is the case. Reserpine-induced
depression has multiple determinants that emerge at different times following initial treatment. Moreover, the impairment appears to be a downstream consequence of reserpine's
effect on the rnonoamines, rather than a direct result of their depletion per se.
An early component of the deficit is evident one hour after drug injection and persists
for about 24 hours. The receptor antagonist for the proinflammatory cytokine IL-1I~ does
not reverse the early deficit. Indeed, this deficit appears too quickly for significant
cytokine expression to occur (Dunn & Swiergiel, 1998; Dunn, Young, Hall, & McNulty,
2002; Fleshner et al., 1995; Nathan, 1987) or the induction of the types of processes that
might lead to significant immune activation (Basu et al., 2002). In this regard, the early
deficit appears to match the time course for catecholamine depletion (Bean et al., 1989).
Given the link between early deficit and adenosine signaling, the process of depleting brain
catecholamines with a single large dose of reserpine may challenge energy homeostasis,
leading to a compensatory adenosine response, and ultimately to behavioral depression.
Late-component depression emerges about 48 hours after reserpine treatment, persists
for at least 24 hours, and then recovers within a week. This deficit is reversed by the

11. CYTOKINE-PURINE INTERACTIONS IN BEHAVIORAL DEPRESSION
1 HOUR POST IL-I~ TREATMENT
15
'-
12
~
0
z
I.-0
_J
199
9
E
6
Z
W
~
3
sVeh
IL-I~
IL-I~
IL-11~
IL-I~
IL-I~
+cVeh
+Caf
+DMSO
+CSC
Fro. 7. Effects of the selective A2A antagonist CSC on swim performance 1 hour after central infusion of IL1~. Rats initially were implated with guide cannula in the right lateral ventricle one week after surgery. One
group received an icy infusion of saline vehicle (sVeh) and 5 groups received an icv infusion of 2 ng 1L-II3.
Forty-five minutes later one IL-113 group received an ip injection of the nonselective adenosine receptor
antagonist caffeine (7 mg/kg) and one IL-I_ group received an ip injection of caffeine vehicle. Other I L - 1
groups received an ip injection of the highly selective A2A antagonist CSC (1.0 mg/kg) or DMSO vehicle.
Swim testing occurred 15 minutes later.
receptor antagonist for IL-1 ~ and is mimicked by direct infusion of the proinflammatory
cytokine into the cerebral ventricles.
Perhaps most important is the finding that both early and late components of the deficit
are reversed by systemic administration of selective A2A receptor antagonists. Although
there is only a small literature on the potential interactions between adenosine and IL-1 ~l, it
is clear that these pathways interact. For instance, application of endotoxin or IL-115 on
PC12 or THP-1 cell upregulates the density of A2A receptors and increases extracellular
concentrations of adenosine (Bshesh et al., 2002; Trincevelli et al., 2002). Several investigators have argued based on such data that adenosine (an anti-inflammatory agent) is
induced as negative feedback on proinflammatory cytokines.
Adenosine A2A receptors also have a distinct and limited distribution in the brain. These
receptors are largely confined to the olfactory epithelium and on the medium spiny neuron
of the striopallidal tract in the striatum (El Yacoubi et al., 2001; Ferre, Fredholm, Morelli,
Popoli, Fuxe, 1997; Svenningsson, LeMoine, Fisone, & Fredholm, 1999). The striatum is a
crucial structure for the integration of sensory, motor, and motivational information. If
behavioral depression can be thought of as the uncoupling of motivation from ongoing
motor activity, then the striatum would seem to be the ideal structure in which such a
process occurs. Thus, activation of Striatal adenosine A2A receptors may act as the switch
for behavioral depression during sickness, traumatic stress, or major depression.

12. 200
MINOR, HUANG, AND FOLEY
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