Roundabout receptor 2 maintains inhibitory control of the adult midbrain
- University of Washington, United States
- Boston University Medical Center, United States
Figures
Figure 1 with 3 supplements
Robo2 VTA mutants have a reduction in the frequency of sIPSCs.
(A) Robo2 is expressed in the VTA, predominantly in dopamine neurons (TH+ cells, arrows), but also non-dopamine neurons (TH- cells, arrowheads). Scale bar = 100 µm (Robo2), 10 µm (merge/zoom). (B) …
Figure 1—figure supplement 1
Robo2 VTA mutants.
(A) In situ hybridization for Robo2 in the rostral, medial, and caudal VTA (image credit: Allen Institute). Scale bar = 500 µm. (B) Traces of the viral transduction of AAV1-Cre-GFP in Robo2lox/lox …
Figure 1—figure supplement 2
Robo2 VTA mutants have normal sEPSCs.
(A) Representative trace from a recorded Ih positive cell. (B) sEPSC representative traces for control and mutant Ih positive cell. (C) sEPSC frequency (left) and cumulative distribution plot …
Figure 1—figure supplement 3
Robo2 VTA mutants.
(A) Representative examples of asymmetrical and symmetrical synapses in control and mutant mice. Blue shading indicates presynaptic terminal. Pink shading indicates postsynaptic terminal. Scale bar =…
Figure 2 with 1 supplement
Robo2 VTA mutants show increased burst activity.
(A,B) Firing rate histogram of a representative cell displaying inhibition to quinpirole that is reversed with eticlopride in control (A) and mutant animal (B). (Left inset) Waveform of represented …
Figure 2—figure supplement 1
in vivo electrophysiology.
(A) Schematic showing tetrode placement in controls and mutants. (B) Representative image showing placement of tetrode in the VTA for in vivo electrophysiological recordings. (C) The distribution of …
Figure 3 with 1 supplement
Robo2 VTA mutants have increased phasic dopamine release.
(A) Schematic of experimental design showing stimulation of the PPTg and FSCV recording in the NAc. (B) Representative voltammogram of dopamine recorded in the NAc in control (left) and mutant …
Figure 3—figure supplement 1
Similar dopamine levels were recorded after MFB stimulation or microdialysis.
(A) Schematic showing the recording electrode sites in the controls and mutants. (B) Representative image showing recording electrode placement in the NAc. (C) Representative image showing …
Figure 4 with 2 supplements
Behavioral characterization of Robo2 VTA mutants show phenotypes in locomotion and cocaine sensitization.
(A) Locomotion measured across three consecutive days and nights showing mutants (n = 31) are hyperactive relative to controls (n = 36). Two-way repeated measures ANOVA, genotype x time interaction, …
Figure 4—figure supplement 1
Robo2 VTA mutants have altered gait.
(A) Hindleg stride length in controls (n = 8 animals) and mutants (n = 8 animals). (B) Foreleg stride length. (C) Hindleg base. (D) Foreleg base, unpaired t-test, t(14) = 2.469, *p<0.05. (E) Print …
Figure 4—figure supplement 2
Additional pharmacological studies on Robo2 VTA mutant mice.
(A) Conditioned place preference (CPP) to cocaine (15 mg/kg) in controls (n = 11) and mutants (n = 10) showed no difference between genotypes. (B) Locomotor response to saline in controls (n = 28) …
Figure 5 with 1 supplement
Adult or embryonic inactivation of Robo2 in dopamine neurons does not recapitulate the Robo2 VTA mutant phenotypes.
(A) Schematic showing expression of tamoxifen-inducible Cre specifically in adult dopamine neurons. Representative images from Slc6a3-iCreERT2 crossed to a Cre-dependent reporter line, Gt(ROSA)26Sort…
Figure 5—figure supplement 1
Embryonic or adult inactivation of Robo2 does not recapitulate the Robo2 VTA mutant phenotypes.
(A, B) Locomotor response to cocaine (20 mg/kg) in controls (n = 12) and mutants (n = 12) on day 1 (A) and day 5 (B). (C) Normalized locomotor response to cocaine by subtracting 60 min pre-cocaine …
Figure 6
Midbrain GABA neuron inactivation recapitulates Robo2 VTA mutant phenotypes.
(A) Schematic of strategy for injecting AAV1-GFP-TetX into the midbrain of Vgat-Cre animals (left). Schematic of Cre-dependent AAV1-GFP-TetX (top). Representative image (bottom) showing viral …
