1. The study aims to map the projections from the nucleus accumbens (NAc) to the ventral tegmental area (VTA) and determine if they are from direct or indirect pathway medium spiny neurons (MSNs). It also aims to map this circuit beyond the midbrain to locate locomotor regions targeted by the NAc. 2. Mice will be injected with viruses to label the D1 and D2 MSN pathways from the NAc to the VTA using GFP. Their brains will then be sliced and visualized under a microscope. 3. Current evidence is unclear whether NAc projections to the VTA are exclusively from D1 or D2 MSNs. Mapping this circuit could classify it

1. Methods
Background: The Classical Basal Ganglia Circuits
An Analogous Circuit
Objective
• The proposed study aims to complete the
following:
1. Illuminate the topography of the
NAcVTA projections and
determine whether they are
reminiscent of direct-pathway
MSNs, indirect-pathway MSNs, or
both.
2. Map the circuitry of this pathway
beyond the ventral striatum and
midbrain, in attempt to show that
the accumbens targets locomotor
regions in the brainstem.
• C57 mice will be injected with Adeno-
associated viruses to label the D1 and D2
tegmental-accumbens pathways using
green fluorescent protein (GFP).
• Similarly, accumbal efferents will be
labeled in a separate cohort of mice.
• These mice will be perfused and their
brains will be sliced sagittally using a
vibratome. Following slicing, the brains will
be mounted and stained on microscope
slides so that the labeling discussed above
can be visualized.
• All experiments will be approved by the
New York State Psychiatric Institute Animal
Care and Use Committee in accordance with
the standards set by the National Institutes
of Health Office of Laboratory Animal
Welfare.
Cortex GLU
Str MSNs
(D2Rs)
GABA GPe GABA
GPi/ EN,
SNr
GABA Thalamus GLU
Motor Cortex
INHIBITED
Cortex GLU Str MSNs (D1Rs) GABA GPi/EN, SNr GABA Thalamus GLU
Motor Cortex
EXCITED
GABA
DA
VTA, SNc Interneurons
ACh
VTA, SNc
DA
Interneurons
ACh
Bridge Collaterals
Direct Striatonigral Pathway
Indirect Striatopallidal Pathway
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• The medium spiny neurons (MSNs) of
the nucleus accumbens (NAc) are a major
component of a “limbic basal ganglia”
circuit. This circuit contains multiple sub-
circuits, one of which consists of
reciprocal projections from the nucleus
accumbens (NAc) to the ventral tegmental
area (VTA) and back.
• While it has been established that the
NAc projects directly to the VTA, the
nature of these projections is
controversial. Xia et al., found that NAc
projects onto GABAergic VTA cells only.
Bocklisch et al. claim that the VTA-
projecting accumbal MSNs consist only of
the dopamine D1 receptor- (D1R-)
expressing sub-type. However, Matsui et
al. recently implied a D2-expressing
projection from NAc to the VTA by
suppressing said pathway via stimulation
of D2Rs.
NAc MSNs VTA
Uchida et al. (2012):
Xia et al. (2011):
NAc MSNs
GABAergic
VTA cells
only
NAc D1R
MSNs
GABA- and
DAergic VTA
cells
Bocklisch et al. (2013):
Matsui et al. (2014):
NAc
GABA- and
DAergic VTA
cells
• This proposed study is the beginning of a
multi-part study aimed at decoding the
accumbal-tegmental pathway and its
relation to facilitation of movement .
• From current evidence, it cannot be
asserted with certainty that the MSNs that
project from the NAc to the VTA are
exclusively D1R- or D2R- expressing. It
seems clear that these projection neurons
express D1Rs, but some may also express
D2Rs. If the nature of these projections
could be clarified, this circuit could be
classified as direct and/or indirect.
• Mapping the topography of this circuit
beyond the striatum and ventral midbrain
could provide insight into treatments for
brain disorders that affect this circuitry
and further establish the NAc as a bridge
between the limbic and motor systems.
• Two thoroughly researched basal ganglia circuits, the direct striatonigral pathway and the
indirect striatopallidal pathway, interact to facilitate desired and inhibit unwanted
behavior. When this delicate balance is disrupted, brain disorders such as Parkinson’s
disease, obsessive-compulsive disorder, and Schizophrenia may result.
Characterizing the Targets of the NAc
• Research exploring subsequent accumbal
projections in this circuit (4) is sparse.
• It has been shown that there are basal
ganglia areas that synapse on the
mesencephalic locomotor region (MLR),
which, in mammals, encompasses the
cuneiform nucleus (CnF) and the
pedunculopontine tegmental nucleus (PPTg).
• The MLR sends glutamatergic afferents to
reticulospinal cells (RS), which activate
central pattern generators (CPGs) for
locomotion in the spinal cord.
• The nucleus accumbens is considered a
bridge between the emotional limbic system
and the motor system. It has been suggested
that there are accumbal projections to the
phylogenically old brainstem regions
discussed above. This could underlie the
changes in motivation and approach behavior
that manifest when this circuit degenerates.
NAc VTA
MSNs (GABA)
?
1
2
2
3
4
Hypothesized Circuit
References
Summary
1 2
3
4
5 6
Legend:
NAc= nucleus
accumbens
VTA= ventral
tegmental area
MLR= mesencephalic
locomotor region
(includes cuneiform
nucleus, CnF and
pedunculopontine
tegmental nucleus
(PPTg)
RF= reticular
formation (contains
reticulospinal, RS,
cells)
CPGs= central
program generators
of locomotion,
located in the spinal
cord