Science - 31 January 2020

and J). The combination of both drugs, how-
ever, resulted in a high density of taD2-SPNs
and a low density of taD1-SPNs throughout
the posterior striatum, indicating that prior
injection of RAC prevented the effects of GBR
on D1-SPN nuclear activity (Fig. 3K). This large-
scale D2-to-D1 suppression was reproduced
when RAC was combined with a full D1 agonist
(fig. S5). Moreover, the distribution of taD2
and taD1-SPN peak density areas tended to
occupy spatially distinct regions of the stria-
tum in all groups, regardless of the pharmaco-
logical cocktail received (Fig. 3, H to K, lower
panels). This opposition was supported by a
strong overall treatment × neuron interaction
(Fig. 3L and table S3). Paradoxically, animals
with D2-dominated (and D1-inhibited) SPN
signaling(Fig.3K)showedunalteredGBR-
induced hyperlocomotion (Fig. 3G), again
suggesting that nuclear activity in SPNs can
be dissociated from behavioral performance
(compare Fig. 1E).

Broad connectivity of the local striatal

network supports an extensive D2- to

D1-SPN transmodulation

We reasoned that the local network in the stria-

tum may reflect large-scale connectivity biases

consistent with the magnitude of the D2-to-D1

modulation reported above. To investigate this,

we used a quantitative network-level approach

to measure connectivity biases using unilat-

eral injections of the herpes simplex virus 1

(HSV1) H129tdTomatoin the posterior striatum

ofdrd2-eGFPmice(Fig.4A).Becausethisvirus

moves along synaptically connected neurons

in the anterograde direction ( 11 ), we mapped

transduced tdTomato+SPNs across large ter-

ritories to reveal the broad connectivity pat-

terns established within the striatum (Fig. 4B

and fig. S6A). Only ~20% of the transduced

SPNs were D2-SPNs, whereas up to ~80% were

D1-SPNs (Fig. 4, C to E, and table S4). These

numbers were similar across all areas of in-

fection, regardless of whether they were ipsi-

or contralateral to the initial injection (fig. S6,

BtoD,andtableS4).Injectionofthevirusin

the prelimbic cortex (i.e., one upstream syn-

apse) provided very similar weights of trans-

duced D1- (~80%) and D2- (~20%) SPNs (fig.

S6, E to G, and table S4).

To confirm that the enhanced connectivity

in D1-SPNs was influenced by a D2-to-D1 drive,

we infected the striatum with HSV1 H129Floxed

virus, an anterograde tracing approach in which

transsynaptic labeling switches from green to

red in the presence of Cre ( 12 )(Fig.4Fandfig.

S7,AtoC).Thismethodallowedustoquantify

cross-system connectivitybyassessingthepro-

portion of (non–Cre-expressing) neurons that

received dual green and red infection within a

critical temporal window (Fig. 4G). Intrastriatal

injection inadora2a-Cre mice revealed equal

proportions of single (tdTom)–and double

(tdTom + eGFP)–labeled SPNs (Fig. 4, H and I,

and table S4). By contrast, infection indrd1a-

Cre mice produced a much lower proportion

Matamaleset al.,Science 367 , 549–555 (2020) 31 January 2020 4of7

103 px

1.5

Beam breaks

Beam breaks (x10

3 )

ACLB

DEF G

HIJ K

P-H3

+

drd2

• eGFP

P-H3

nuclei+

(in D1-SPNs)

P-H3

+ nuclei

(in D2-SPNs)

D2-SPNs D1-SPNs D2-SPNs D1-SPNs D2-SPNs D1-SPNs D2-SPNs D1-SPNs

1 st Inj

01530

2 nd Inj Perfusion

Raclopride

GBR12783

Vehicle

Groups

Raclopride GBR12783

n.s.

Open field Open field

L

D

Veh+Veh

RAC+Veh

Veh+GBR

RAC+GBR

(Veh + Veh) (Rac + Veh) (Veh + GBR) (RAC + GBR)

100 μm

0.0

0.0

1.0

2.0

3.0

0

200

400

600

800

1.5

0.0

1.5

0.0

1.5

0.0

01530 (min) 01530 (min) 01530 (min)^01530 (min)

0.0

0.5

1.0

1.5

Fig. 3. Overstimulated SPN systems compete for space in the striatum.
(A) Confocal micrographs showing the effects of D2R blockade (Raclopride,
0.3 mg/kg) and dopamine transporterinhibition (GBR12783, 15 mg/kg)
on taSPNs. (B) Two-injection experimental design applied to each group prior
to perfusion (eight mice per group). Ambulation was measured in an open
field. (C) Ambulation (beam brakes) recorded after the second injection
(min 15 to 30). (DtoG) Ambulatory activity per minute. Right: ambulatory
trajectory (start: blue; finish: red) in one example mouse after the second

injection. (HtoK) Top: maps of taD2- and taD1-SPNs in the striatum of an

example mouse. Bottom: distribution contour plots delimitating regions of

increasing P-H3+nuclear density in D2- (left) and D1- (right) SPN systems

separately. Isodensity curves are pseudocolored from low (blue) to high

(red) relative densities (31,542 taD1-SPNs and 29,100 taD2-SPNs mapped).

(L) Quantification of P-H3+nuclei (counts × 10^3 ) distributed in D2- and

D1-SPNs in each group (9 to 12 sections per group). *, significant overall/

simple effect (black) and interaction (red). n.s., not significant (table S3).

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