aav dj cmv mcherry t2a icre  (Vector Biolabs)

Journal: Cell

Article Title: A Neuro-hormonal Circuit for Paternal Behavior Controlled by a Hypothalamic Network Oscillation

doi: 10.1016/j.cell.2020.07.007

Figure Lengend Snippet: Following Sexual Experience, Prl Is Sufficient and Necessary for Paternal Behavior (A) Schematic of the experimental design used to determine whether systemic injection of Prl raises serum Prl levels and alters pSTAT5 immunofluorescence in the MPOA. (B) Representative confocal images and quantification of pSTAT5 immunofluorescence in the MPOA in saline-injected (top) and Prl-injected (bottom) adult virgin male rats. aca, anterior commissure. (C) Quantification of pSTAT5 cells in the MPOA in control and Prl-injected rats (n = 24 MPOA sections per group from 5 rats per group, two-tailed unpaired t test, p < 0.0001 between saline and Prl). (D) Quantification of serum Prl in control and Prl-injected rats (n = 5 per group, two-tailed unpaired t test, p < 0.0001 between saline and Prl). (E) Schematic of the experimental design used to assess whether i.p. administration of Prl can induce paternal behavior in sexually experienced rat sires. (F) Representative behavior raster plots of a rat sire in the pup retrieval test after saline (top) and Prl (bottom) injection. (G–L) Quantification of indices of paternal investment in rat sires (n = 13 and 16 trials with saline and Prl i.p. injection, respectively, one-way repeated-measures ANOVA with Dunn’s post hoc test). Note the improved performance on most parameters in response to Prl administration. (G) Pup retrieval duration; p = 0.0774 between saline and Prl. (H) Time spent in proximity to pups; p = 0.0001 between saline and Prl. (I) Crouching duration; p = 0.0003 between saline and Prl. (J) Pup grooming duration; p = 0.0012 between saline and Prl. (K) Mouthing duration; p = 0.0007 between saline and Prl. (L) Nesting duration; p = 0.0923 between saline and Prl. (M) Schematic of the experimental design used to determine the effect of conditional knockout of the Prl-R on pSTAT5 immunoreactivity in the MPOA. (N) Representative bright-field images and quantification of pSTAT5 immunoreactivity in the MPOA in AAV-mCherry-injected (control) and AAV-Cre-injected (Prl-R knockout) adult male mice. (O) Quantification of pSTAT5 cells in the MPOA in AAV-mCherry- and AAV-Cre-injected mice (n = 6 bilateral MPOA sections per group from 6 mice per group, two-tailed unpaired t test, p < 0.0001 between AAV-mCherry and AAV-Cre). (P) Schematic of the experimental design used to assess whether MPOA Prl-R knockout in mouse sires influences paternal behavior. (Q) Representative behavior raster plots of two individual mouse sires in the pup retrieval test after AAV-mCherry (top) and AAV-Cre (bottom) injection, respectively, in the MPOA. (R–W) Quantification of indices of paternal investment in mouse sires (n = 6 trials/mice/group with AAV-mCherry and AAV-Cre MPOA injection, respectively; two-sided Mann-Whitney U test was used for all comparisons). (R) Duration of pup retrieval behavior exhibited by mouse sires (AAV-mCherry versus AAV-Cre, p = 0.0195). (S) Time spent in proximity to pups (AAV-mCherry versus AAV-Cre, p = 0.0022). (T) Duration of crouching above pups (AAV-mCherry versus AAV-Cre, p = 0.0130). (U) Duration of pup grooming behavior exhibited by mouse sires toward pups (AAV-mCherry versus AAV-Cre, p = 0.0346). (V) Total number of pups retrieved at the end of the test (maximum number of pups = 4, AAV-mCherry versus AAV-Cre, p = 0.0152). (W) Duration of nesting behavior exhibited by mouse sires (AAV-mCherry versus AAV-Cre, p = 0.0931). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. Each dot represents a readout from a single session in the pup retrieval test. Duplicates or triplicates were collected and are presented as single dots for each animal. Violin plots indicate the non-normal distribution of behaviors. All experiments were performed in male rodents. See also , , and .

Article Snippet: AAV/DJ-CMV-mCherry-T2A-iCre , Vector Biolabs , Cat# VB7600.

Techniques: Injection, Immunofluorescence, Two Tailed Test, Knock-Out, MANN-WHITNEY

Journal: Cell

Article Title: A Neuro-hormonal Circuit for Paternal Behavior Controlled by a Hypothalamic Network Oscillation

doi: 10.1016/j.cell.2020.07.007

Figure Lengend Snippet: Validation of Prl-R Gene Deletion, Related to Figure 6 (A) Schematic of the experimental design used to determine if stereotactic injection of AAV-Cre in the MPOA of prolactin-receptor floxed mice leads to deletion of the prolactin receptor gene. As Cre-mediated deletion of the prolactin gene involves simultaneous inversion of the prolactin receptor gene and eGFP expression, eGFP fluorescence can serve as a marker of the injection site and successful recombination. (B) Representative brightfield images and quantification of eGFP immunoreactivity in the MPOA, in AAV-mCherry (control) and AAV-Cre (prolactin receptor knockout) injected adult male mice. oc, optic chiasm; 3V, third ventricle. Note the absence of eGFP immunoreactivity in the control (AAV-mCherry injected) mice, in contrast to the expression of eGFP (a marker of the prolactin receptor gene deletion) in the MPOA of AAV-Cre injected mice. (C) Quantification of eGFP cells in the MPOA in AAV-mCherry and AAV-Cre-injected mice (n = 6 bilateral MPOA sections per group from 6 mice per group, two-sided Mann–Whitney U test, p = 0.0022 between AAV-mCherry and AAV-Cre). ∗∗ p < 0.01. In box-and-whisker plots, center lines indicate medians, box edges represent the interquartile range, and whiskers extend to the minimal and maximal values. All experiments presented in Figure S5 were performed in male mice.

Article Snippet: AAV/DJ-CMV-mCherry-T2A-iCre , Vector Biolabs , Cat# VB7600.

Techniques: Injection, Expressing, Fluorescence, Marker, Knock-Out, MANN-WHITNEY, Whisker Assay

Journal: Cell

Article Title: A Neuro-hormonal Circuit for Paternal Behavior Controlled by a Hypothalamic Network Oscillation

doi: 10.1016/j.cell.2020.07.007

Figure Lengend Snippet:

Article Snippet: AAV/DJ-CMV-mCherry-T2A-iCre , Vector Biolabs , Cat# VB7600.

Techniques: Plasmid Preparation, Recombinant, Enzyme-linked Immunosorbent Assay, Competitive ELISA, Software