Article Figures & Data
Figures
- Figure 1.
Visualization of Drd2-Pet1 serotonergic neurons and the loss of Drd2 gene expression in Drd2Pet1-CKO mice. A, Drd2-Pet1 neurons are intersectionally labeled with GFP (green) and Pet1-only positive cell bodies labeled with mCherry (magenta) in a coronal brain section of the DR from a P90 triple transgenic Drd2-Cre;Pet1-Flpe;RC:FrePe mouse. Scale bars: 200 μm. B, Intersectional genetic strategy: expression of Drd2-Cre and Pet1-Flpe transgenes results in dual recombination of intersectional allele, RC:FrePe, labeling cells expressing Drd2 and Pet1 with GFP. C, Dual immunohistochemistry for GFP (green) and 5-HT (serotonin, magenta) coupled with FISH detection of Drd2 mRNA, which shows co-localization of intersectionally labeled Drd2-Pet1 neuron cell bodies with 5-HT and Drd2 mRNA. Scale bars: 10 μm. D, Strategy for conditional deletion of Drd2 in serotonergic neurons (referred to throughout as Drd2Pet1-CKO). Cre recombination excises Drd2 exon 2 (magenta) producing serotonergic-specific (boxed in green) deletion of Drd2 gene sequences. E, Percentage (mean ± SEM) of Pet1+ serotonergic neurons that express Drd2 in control (n = 6) versus Drd2Pet1-CKO (n = 6) shows reduction of Drd2 expression in Pet1+ neurons (controls: 15.23 ± 2.41 Drd2-Pet1 dual positive neurons per brain, Drd2Pet1-CKO: 3.87 ± 0.73 Drd2-Pet1 dual positive neurons per brain, p = 0.0011, unpaired t test). Filled black diamonds represent male mice, open gray circles represent female mice. F, G, FISH on (F) control and (G) Drd2Pet1-CKO tissue. Drd2 transcripts detected in Pet1+ cells in control sections, but not in Drd2Pet1-CKO mice, indicative of loss of Drd2. cre transcript is not present in control (F, far right) but is present in Drd2Pet1-CKO Pet1 cells, as expected (G, far right). Pet1, Drd2, and cre transcript are shown separately in grayscale. Note Drd2 expression remains in non-Pet1 cells (arrow). Dotted lines drawn to encircle DAPI nuclei. Scale bars: 25 μm.
- Figure 2.
Drd2Pet1-CKO mice are largely behaviorally normal. Drd2Pet1-CKO (blue symbols) mice show behaviors indistinguishable from controls (black symbols) in measures of locomotion: (A, B) open field test and (C) rotarod; measures of anxiety-like and depression-like behavior: (D) elevated plus maze, (E) tail suspension test, and (F) forced swim test; or learning and memory: (G) contextual fear conditioning and (H) water T maze; n = 15 control mice (8 males, 7 females) and 11 Drd2Pet1-CKO (6 males, 5 females), except for C where, n = 21 control mice (14 males, 7 females) and 18 Drd2Pet1-CKO (13 males, 5 females). Each symbol represents one animal, error bars represent SEM. No significant differences (p > 0.05) between Drd2Pet1-CKO and controls were observed. No sex-specific (male vs female) phenotypes observed. For assay details, see Materials and Methods; for statistical details, see Table 2.
- Figure 3.
Drd2Pet1-CKO females, but not males, display attenuated acoustic startle responses (ASR). A, Schematic of ASR experimental design. After an initial 5-min acclimation, mice are exposed to 10 blocks of 11 trials of auditory stimuli ranging from 20 to 120 dB in quasi-randomized order with a 10- to 20-s intertrial interval (ITI). B, Schematic of ASR measurement apparatus, mouse is placed in a perforated holding chamber atop transducer platform adjacent to speaker (for detailed description, see Materials and Methods). C, F, Averaged ASR magnitudes (mean ± SEM) across increasing stimulus intensities in (C) male Drd2Pet1-CKO (blue, n = 13) and controls (black, n = 14), no significant difference, p = 0.7745, two-way ANOVA and (F) female Drd2Pet1-CKO (blue, n = 15) and controls (black, n = 16), Drd2Pet1-CKO females display significantly attenuated ASR, p = 0.0011, two-way ANOVA. D, G, Group averaged ASR for 10 trials at 110-dB stimulus in (D) males and (G) females, demonstrates no habituation to the startle stimulus; x-axis numbers refers to trial number out of 110 total trials. E, H, No significant differences in latency to startle are observed in (E) males, p = 0.1319, two-way ANOVA and (H) females, p = 0.5452, two-way ANOVA. I, J, No significant differences in prepulse inhibition of acoustic startle are observed in (I) males (n = 8 control, 6 Drd2Pet1-CKO), p = 0.4325, two-way ANOVA or (J) females (n = 7 control, 5 Drd2Pet1-CKO, p = 0.4380, two-way ANOVA).
- Figure 4.
Drd2Pet1-CKO mice show normal auditory responses. A, B, Average ABR waveforms at 16 kHz for (A) control (black, n = 10) and Drd2Pet1-CKO (blue, n = 7) males and (B) for control (black, n = 8) and Drd2Pet1-CKO (blue, n = 7) females. Average is shown by darker lines and shaded area shows SEM. C, F, ABR amplitudes for control (black) and Drd2Pet1-CKO (blue; C) male and (F) female mice for ABR peaks 1 through 3. No significant difference was observed between control and Drd2Pet1-CKO: males, p = 0.2032; females, p = 0.1387, two-way ANOVA. D, F, Latencies for control (black) and Drd2Pet1-CKO (blue; D) male and (G) female mice for ABR peaks 1 through 3. No significant difference was observed between control and Drd2Pet1-CKO: males, p = 0.0804; females, p = 0.9430, two-way ANOVA. Amplitudes and latencies shown at 80-dB SPL. E, H, ABR thresholds for control (black) and Drd2Pet1-CKO (blue; E) male and (H) female mice across frequencies tested (5.6, 8, 16, and 32 kHz). No significant difference was observed between control and Drd2Pet1-CKO mice, p > 0.05 at all frequencies, unpaired t test.
- Figure 5.
Drd2Pet1-CKO males, but not females, display increased social dominance. A, B, Three chambered social interaction assay. No significant difference in time spent investigating a stranger mouse or an empty holder for (A) males (n = 8 controls compared with 6 Drd2Pet1-CKO, p = 0.541, unpaired t test) and (B) females (n = 7 controls compared with n = 5 Drd2Pet1-CKO, p = 0.358, unpaired t test). Investigation time is binned into 5-min intervals where white bars indicate first 5 min of assay and colored bars indicate last 5 min of assay. As expected, mice of both genotypes spent significantly less time investigating the empty holder than the stranger mouse noting that females of both genotypes only did so during the first 5 min of the assay. C, Resident intruder assay of aggression. No significant difference in the average attack bites per day delivered to a Swiss Webster intruder mouse was observed between Drd2Pet1-CKO males (n = 26, 4.07 ± 1.50 bites) aggression levels were not significantly different from controls (n = 24, 1.77 ± 0.39 bites; Mann–Whitney, two-tailed, U = 289.5, p = 0.6649). D, Schematic of tube test (for details of assay, see Materials and Methods). Schematic created with BioRender. E, Drd2Pet1-CKO males (n = 24) demonstrate more dominance behavior than controls (n = 24) as they displayed increased winning in the tube test (controls: 34.17 ± 9% wins, Drd2Pet1-CKO: 65.83 ± 9% wins, p = 0.0065, Mann–Whitney, two-tailed, U = 166). Female Drd2Pet1-CKO (n = 23) showed no difference in social dominance compared with controls (n = 23; controls: 51.3 ± 8%, Drd2Pet1-CKO: 48.7 ± 8% wins, p = 0.8123 Mann–Whitney, two-tailed, U = 253).
- Figure 6.
Sex-specific transcript level differences in Drd2-Pet1 neurons. A, Dual immunohistochemistry and FISH depicting green GFP+ Drd2-Pet1 neurons along with transcript puncta in male (top) and female (bottom) brain sections from Drd2-Cre;Pet1-Flpe;RC-FrePe mice. Drd2 (cyan) and Gad2 (magenta) expression shown together and separately in gray scale. Scale bar: 10 μm. B, Number of Drd2-Pet1 neurons (GFP-positive cells in Drd2-Cre;Pet1-Flpe;RC-FrePE mice) per animal in males (black diamonds, n = 7) and females (open gray circles, n = 7) is not significantly different. Males: 410.40 ± 55.30 cells/brain, females: 313 ± 87.52 cells/brain, p = 0.4336, unpaired t test. C, Drd2-Pet1 neuron soma size (GFP+ cell body) does not differ in males (n = 5 males) versus females (n = 5 females), p = 0.3372, unpaired t test. D, Number of FISH mRNA puncta per cell in males versus females. Male cells have significantly more Gad2 puncta than female cells [20.46 ± 2.243 in males (n = 5) vs 12.20 ± 2.427 in females (n = 5), p = 0.0370, unpaired t test]. E, 86.47 ± 4.181% of male Drd2-Pet1 cells express Gad2 versus female 74.00 ± 5.168% in female cells, p = 0.0975, unpaired t test. Error bars indicate SEM throughout. For C, D, larger symbols outlined in black represent animal averages used for statistical analysis, smaller symbols represent individual cells, matched in color to the average.
- Figure 7.
Gad2 expression in Drd2Pet1-CKO cells. A, FISH with probes to Drd2 exon 7/8 (D2-E7/8, green) and Drd2 exon 2 (D2-E2, magenta) in Pet1 (white) cells in control (top) and Drd2Pet1-CKO (bottom) DR tissue. D2-E7/8, D2-E2, and Pet1 expression shown together and separately in gray scale. B, Percent of Pet1+ cells (left and middle) with Drd2-Exon7/8 expression in control (35.97 ± 2.403%, n = 6) and Drd2Pet1-CKO (36.53 ± 3.621%, n = 6), p = 0.8998, unpaired t test. Data also shown for percent of cre cells (right) with Drd2-Exon7/8, 34.91 ± 2.238%, compared with Pet1 probe control and Drd2Pet1-CKO p = 0.9051, one-way ANOVA. Males, black diamonds, females, open gray circles. C, FISH showing cre+ Drd2Pet1-CKO cells (white) with Drd2-Exon7/8 (green) and Gad2 (red) in male (top) and female (bottom) in the DR nucleus. Drd2-Exon7/8, Gad2, and cre are shown together and separately in gray scale. Scale bar: 10 μm. D, A larger percentage of male Drd2Pet1-CKO cells (87.44 ± 3.034%) express Gad2 versus female Drd2Pet1-CKO cells (75.76 ± 0.5862%), *p = 0.0157, unpaired t test. E, Number of Gad2 mRNA puncta per cell in Drd2Pet1-CKO cells in males (n = 6) versus females (n = 4). Male cells have 14.25 ± 1.325 Gad2 puncta per cell compared with 10.13 ± 2.074 in female cells, p = 0.1151, unpaired t test. F, Drd2Pet1-CKO nucleus size (area used to quantify puncta levels) does not differ in males (n = 6 males) versus females (n = 4 females), p = 0.3497, unpaired t test. Error bars indicate SEM throughout. For E, F, larger symbols outlined in black represent animal averages used for statistical analysis, smaller symbols represent individual cells, matched in color to the average.
- Figure 8.
Drd2-Pet1 neuron electrophysiological properties in male versus female mice. Membrane and AP characteristics were analyzed in GFP-marked Drd2-Pet1 male and female neurons using whole-cell patch-clamp electrophysiology in acute brain slices from triple transgenic Drd2-Cre;Pet1-Flpe;RC-FrePe mice. Membrane potential (A), membrane resistance (B), AP threshold (C), AP amplitude (D), and AHP amplitude (F) do not differ in male (n = 19) or female (n = 44) Drd2-Pet1 neurons while (E) male Drd2-Pet1 neurons had a significantly longer (2.847 ± 0.155 ms, n = 19 cells) AP duration than in females (2.54 ± 0.094 ms, n = 44 cells, p = 0.0275, unpaired t test).
- Figure 9.
Drd2-Pet1 neuron axon terminals target brain regions involved in sensory processing and defensive behavior in both male and female mice. A, Intersectional genetic strategy: expression of Drd2-Cre and Pet1-Flpe transgenes results in dual recombination of intersectional allele, RC-FPSit, to label boutons of Drd2-Pet1 neurons with Synaptophysin-GFP. B, Representative images of Drd2-Pet1 boutons in the SOC and IC. GFP+ (green, marked with arrows) boutons co-localize with 5-HT (magenta) staining. DAPI-stained nuclei shown in blue. Scale bar: 25 μm. C, Quantification of the percent target area occupied by projections for all ten brain regions examined (for quantification protocol, see Materials and Methods). Target areas analyzed include brain regions involved in auditory processing and social behavior including the CNC, SOC, LL, IC, PNC, mHb, dLGN, mPOA, and PAG. The DPGi was also examined. No significant differences in projection area innervation were observed between males (n = 5) and females (n = 6). D, Example graph showing correlation between innervation density of auditory brain regions differs in males compared with females. Each dot represents one animal. Values are shown as Pearson’s correlation coefficient (r), and * indicates p < 0.5 in a two-tailed test. E, Pairwise correlations shown for male and female innervation density in auditory brain regions. Heatmaps represent high correlation (green) and low correlation (black) between CNC, SOC, LL, IC, and PNC.
- Figure 10.
GABA immunoreactivity localizes to soma, but not axonal projections, of Drd2-Pet1 neurons. A, GABA staining (magenta) co-localizes with many Drd2-Pet1 neuron soma (green GFP-positive cell bodies in Drd2-Cre;Pet1-Flpe;RC-FPSit mice) in the DR in a punctate manner (top), inset of boxed region showing neuron soma positive for GFP and GABA. Some Drd2-Pet1 neuron soma are immuno-negative for GABA (bottom). Dotted lines encircle GFP-positive cell body. B, C, No GFP-positive Drd2-Pet1 boutons (green) co-localize with GABA staining (magenta) in brain regions examined, shown here, representative images from SOC (B) and IC (C), noting a GABA-positive soma is visible in the image of the IC. D, GABA-positive immunoreactivity in the corpus callosum demonstrating detection of GABA boutons. ChAT (white) staining was used throughout for anatomic localization. Scale bars: 25 μm (left panel) and 10 μm (inset). DAPI-stained nuclei shown in blue.
Tables
Probe S1 S2 Prominence R2 RMSE MAE E2 0.25 1 175 0.8696 0.5957 0.2458 E7/8 0.5 1 100 0.9421 0.8054 0.35 Cre 0.5 1 100 0.9679 3.829 2.2244 Fev 0.25 2 75 0.9555 4.414 3.7047 Gad2 0.25 16 150 0.8568 2.804 1.7973 Summary of settings and performance in a linear regression for semi-automated protocol versus hand counts for each FISH probe.
Behavior/experiment Line Data structure
(normality)Type of test Power Comparison F/df p Validation of Drd2 CKO 1E Yes Unpaired t test Control vs Drd2Pet1-CKO t = 4.514, df = 10 p = 0.0011 Open field distance 2A Yes Repeated-measures ANOVA F1, genotype F(1,24) = 0.6405 p = 0.4314 F2, time F(11,264) = 47.99 p < 0.0001 (F1 × F2) F(11,264) = 0.8441 p = 0.5960 Open field % distance traveled 2B Yes Unpaired t test Control vs Drd2Pet1-CKO t = 1.781, df = 24 p = 0.0876 Rotarod 2C No Mann–Whitney, two-tailed Control vs Drd2Pet1-CKO M–W U = 142 p = 0.1899 Elevated plus maze (% time in open arm) 2D Yes Unpaired t test Control vs Drd2Pet1-CKO t = 1.250, df = 24 p = 0.2234 Tail suspension test 2E Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.3485, df = 24 p = 0.7305 Forced swim test 2F Yes Repeated-measures ANOVA F1, genotype F(1,24) = 0.2678 p = 0.6095 F2, time F(5,120) = 8.916 p < 0.0001 (F1 × F2) F(5,120) = 0.3090 p = 0.9067 Contextual fear conditioning (baseline freezing) 2G Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.6682, df = 24 p = 0.5104 Contextual fear conditioning (test freezing) Unpaired t test Control vs Drd2Pet1-CKO t = 0.0127, df = 24 p = 0.9900 Water T maze (%correct during acquisition) 2H Yes Repeated-measures ANOVA F1, genotype F(1,24) = 0.08249 p = 0.7764 F2, time F(4,89) = 50.12 p < 0.0001 (F1 × F2) F(4,89) = 0.6698 p = 0.6147 Water T maze (%correct during reversal) Yes Repeated-measures ANOVA F1, genotype F(1,24) = 0.1631 p = 0.6899 F2, time F(4,96) = 172.4 p < 0.0001 (F1 × F2) F(4,96) = 1.477 p = 0.2153 ASR (M) 3C Yes Repeated-measures ANOVA F1, genotype F(1,25) = 0.0840 p = 0.7745 F2, dB F(10,250) = 28.99 p < 0.0001 (F1 × F2) F(10,250) = 0.3037 p = 0.9798 ASR habituation (M) 3D Yes Pearson r correlation Control trial number × startle response r = −0.195 p = 0.5893 Drd2Pet1-CKO trial number × startle response r = 0.136 p = 0.7079 ASR latency (M) 3E Yes Repeated-measures ANOVA F1, genotype F(1,25) = 2.425 p = 0.1319 F2, dB F(10,250) = 21.67 p < 0.0001 (F1 × F2) F(10,250) = 0.4722 p = 0.9071 ASR (F) 3F Yes Repeated-measures ANOVA F1, genotype F(1,29) = 13.26 p = 0.0011 F2, dB F(10,29) = 35.29 p < 0.0001 (F1 × F2) F(10,290) = 7.475 p < 0.0001 ASR habituation (F) 3G Yes Pearson r correlation Control trial number × startle response r = 0.1171 p = 0.7473 Drd2Pet1-CKO trial number × startle response r = 0.05165 p = 0.8873 ASR latency (F) 3H Yes Repeated-measures ANOVA F1, genotype F(1,29) = 0.3748 p = 0.5452 F2, dB F(10,290) = 20.59 p < 0.0001 (F1 × F2) F(10,290) = 1.058 p = 0.3953 PPI (M) 3I Yes Repeated-measures ANOVA F1, genotype F(1,12) = 0.6625 p = 0.4315 F2, prepulse dB F(2,24) = 42.86 p < 0.0001 (F1 × F2) F(2,24) = 4.104 p = 0.0293 PPI (F) 3J Yes Repeated-measures ANOVA F1, genotype F(1,10) = 0.6526 p = 0.4380 F2, prepulse dB F(2,20) = 31.34 p < 0.0001 (F1 × F2) F(2,20) = 1.609 p = 0.2249 ABR amplitude (M) 4C Yes Repeated-measures ANOVA F1, genotype F(1,15) = 1.770 p = 0.2032 F2, peak F(2,30) = 59.09 p < 0.0001 (F1 × F2) F(2,30) = 1.059 p = 0.3595 ABR latency (M) 4D Yes Repeated-measures ANOVA F1, genotype F(1,15) = 3.515 p = 0.0804 F2, peak F(2,30) = 1171 p < 0.0001 (F1 × F2) F(2,30) = 3.121 p = 0.0587 ABR threshold (M) 4E 5.6 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.9535, df = 14 p = 0.3565 8 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 1.894, df = 14 p = 0.0791 16 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 1.103, df = 14 p = 0.2887 32 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 2.129, df = 7 p = 0.0708 ABR amplitude (F) 4F Yes Repeated-measures ANOVA F1, genotype F(1,13) = 2.489 p = 0.1387 F2, peak F(2,26) = 72.52 p < 0.0001 (F1 × F2) F(2,26) = 0.0487 p = 0.9525 ABR latency (F) 4G Yes Repeated-measures ANOVA F1, genotype F(1,13) = 0.0053 p = 0.9430 F2, peak F(2,26) = 4360 p < 0.0001 (F1 × F2) F(2,26) = 0.0822 p = 0.9213 ABR threshold (F) 4H 5.6 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.1566, df = 13 p = 0.8770 8 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.1592, df = 14 p = 0.8757 16 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.9600, df = 14 p = 0.3533 32 Yes Unpaired t test Control vs Drd2Pet1-CKO t = 1.644, df = 9 p = 0.1346 Social interaction (M, %time with stranger) 5A Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.6283, df = 12 p = 0.5415 Social interaction (F, %time with stranger) 5B Yes Unpaired t test Control vs Drd2Pet1-CKO t = 0.9598, df = 10 p = 0.3598 Resident-intruder assay 5C No Mann–Whitney, two-tailed Control vs Drd2Pet1-CKO M–W U = 289.5 p = 0.6649 Tube test of social dominance 5E Male No Mann–Whitney, two-tailed Control vs Drd2Pet1-CKO M–W U = 166 p = 0.0065 Female No Mann–Whitney, two-tailed Control vs Drd2Pet1-CKO M–W U = 253 p = 0.8123 Drd2-Pet1 neuron count 6A Yes Unpaired t test Male vs female t = 0.8160, df = 12 p = 0.4304 Soma size 6C Yes Unpaired t test Male vs female t = 1.021, df = 8 p = 0.3372 Gene expression 6D Dmd Yes Unpaired t test Male vs female t = 0.9581, df = 7 p = 0.3699 Drd2 Yes Unpaired t test Male vs female t = 1.514, df = 8 p = 0.1686 Gad2 Yes Unpaired t test Male vs female t = 2.498, df = 8 p = 0.0370 Serpini1 Yes Unpaired t test Male vs female t = 1.459, df = 7 p = 0.1879 %Gad2+ Drd2-Pet1 neurons 6E Yes Unpaired t test Male vs female t = 1.876, df = 8 p = 0.0975 % Drd2-Exon7/8+ 7B Yes Unpaired t test Control vs Drd2Pet1-CKO with Pet1 probe t = 0.1291, df = 10 p = 0.8998 Yes One-way ANOVA Control/Pet1 probe vs Drd2Pet1-CKO/
Pet1 probe vs Drd2Pet1-CKO /Cre probeF(2,19) = 0.1003 p = 0.9051 % Gad2 in Cre+ neurons 7D Yes Unpaired t test Male vs female t = 3.057, df = 8 p = 0.0157 Gad2 punctae per cell 7E Yes Unpaired t test Male vs female t = 1.768, df = 8 p = 0.1151 Nucleus area 7F Yes Unpaired t test Male vs female t = 0.9931, df = 8 p = 0.3497 Resting membrane potential 8A Yes Unpaired t test Male vs female t = 0.2113, df = 61 p = 0.8334 Membrane resistance 8B Yes Unpaired t test Male vs female t = −0.4084, df = 61 p = 0.6844 AP threshold 8B Yes Unpaired t test Male vs female t = 1.8197, df = 61 p = 0.0737 AP amplitude 8D Yes Unpaired t test Male vs female t = −1.0474, df = 61 p = 0.2990 AP duration 8E Yes Unpaired t test Male vs female t = −2.2583, df = 61 p = 0.0275 AHP amplitude 8F Yes Unpaired t test Male vs female t = 1.350, df = 61 p = 0.1821 Innervation densities 9C DPGi Yes Unpaired t test Male vs female t = 1.285, df = 9 p = 0.2308 PAG Yes Unpaired t test Male vs female t = 0.2398, df = 9 p = 0.8158 mPOA Yes Unpaired t test Male vs female t = 0.1978, df = 9 p = 0.8476 DLG Yes Unpaired t test Male vs female t = 0.07798, df = 9 p = 0.9395 mHb Yes Unpaired t test Male vs female t = 0.6732, df = 9 p = 0.5178 PnC Yes Unpaired t test Male vs female t = 0.7901, df = 9 p = 0.4498 IC Yes Unpaired t test Male vs female t = 0.5350, df = 9 p = 0.6056 LL Yes Unpaired t test Male vs female t = 0.9100, df = 9 p = 0.3865 SOC Yes Unpaired t test Male vs female t = 0.9282, df = 9 p = 0.3775 CNC Yes Unpaired t test Male vs female t = 0.2997, df = 9 p = 0.7712 Statistical values are provided for behavioral analyses of Drd2Pet1-CKO mice and comparison of Drd2-Pet1 neuron properties in male versus female mice. Figure numbers are included to reference corresponding graphs. Statistical analyses were conducted in GraphPad Prism version 8.1.
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