Journal: bioRxiv

Article Title: Dopamine in the basal amygdala signals salient somatosensory events during fear learning

doi: 10.1101/716589

Figure Lengend Snippet: VTA neurons, and amongst them dopamine neurons, respond to footshocks and acquire CS - responsiveness. ( A ) Schematic showing an optrode with 16 recording channels implanted in the VTA of a DAT Cre × ChR2-eYFP mouse. ( B ) Post-hoc histological image showing the placement of an optrode (dashed line) in the VTA in the example mouse of panels B – H (FT7612). The tracks of two tetrodes are visible (arrows). Bottom, scheme of the design of four tetrodes (T1 - T4) around the optical fiber. ( C ) Illustration of optogenetic identification of putative DAT + units. Raster plot for four electrodes (E1-E4) of one tetrode, showing unsorted spikes aligned to the onsets of n = 100, 2 ms – long laser light pulses (blue shading). Spikes at 2-8 ms after the light pulse were collected and subjected to spike clustering (see Materials and Methods). ( D ) Time course of freezing of the example mouse during the three days of the fear learning protocol. ( E ) Spiking activity of a single opto-tagged putative DAT + unit during day 2 (training day). AP frequency (top) and z-score (bottom) are shown in response to the CS (averages over the n = 30 tone presentations for each tone block), as well as in response to footshocks (averages over the n = 6 footshock presentations; right). ( F ) Spiking activity of the same unit as (E), but for responses to CS presentations on day 3 (threat memory retrieval). Right, AP-waveforms for the unit shown in E, F. ( G, H ) Responses of all units in this mouse (FT7612) to tone presentations during day1 - day 3 (G), and to footshocks on day 2 (H). Z-scores were analyzed and plotted as a function of time (number of tone block). Units classified as CS - entrained are shown in pink, and the others in gray. Thick red and black lines show average ± S.E.M. across these groups, respectively. Square symbols connected by lines represent DAT + units identified by optotagging. ( I, J, K ) Venn-type diagrams showing the number of units in the different response classes and their overlap. The data in (I) is from the example mouse shown in panels B-H. Note the overall similar distribution and overlap of response types across n = 3 mice. Two US responsive units in mouse FT6963 showed a reduction in firing upon the footshock (J, dashed areas).

Article Snippet: A classical auditory cued threat memory paradigm was performed in a conditioning chamber of a NIR Video Fear Conditioning Optogenetics Package for Mouse (MED-VFC-OPTO-M, Med Associates Inc., VT, USA).

Techniques: Activity Assay, Blocking Assay

Journal: The Journal of Biological Chemistry

Article Title: An Antibody Biosensor Establishes the Activation of the M 1 Muscarinic Acetylcholine Receptor during Learning and Memory * ^♦

doi: 10.1074/jbc.M115.681726

Figure Lengend Snippet: M 1 mAChR is activated in the hippocampus following drug treatment and memory acquisition. A, C57/BL6/NTAC mice were injected (i.p.) with vehicle or xanomeline (5 mg/kg). After 30 min, tissues were fixed by transcardial perfusion, and sections were obtained and stained with the phospho-specific serine 228 antibody and with DAPI stain to reveal the nuclei. Shown are representative sections through the CA1 region of the hippocampus. B, C57/BL6/NTAC mice ( Wild-Type ) or M 1 mAChR-knock-out mice ( M1-KO ) were injected (intraperitoneally) with BQCA (15 mg/kg) or vehicle, and after 30 min hippocampal membranes were prepared from which the M 1 mAChR was immunoprecipitated. The sample was then processed in Western blots, which were probed with phospho-specific serine 228 antibody or an M 1 mAChR-specific antibody to detect total M 1 mAChR. C, quantification of Western blots from B . The data are presented as means ± S.E. ( n = 3). Statistical analysis uses Student's paired t test. D, C57/BL6/NTAC mice were subjected to a fear conditioning training protocol or to an unpaired immediate foot shock as a control; 30 min later tissue was fixed by transcardial perfusion, and sections obtained and stained with phosphorylated Ser 228 -specific antibody ( upper panel ) or anti-c-FOS antibody ( lower panel ). All the data shown are typical of at least three independent experiments.

Article Snippet: Male C57Bl6/NTAC mice (8–15 weeks old) were placed in the conditioning chamber (Stoelting ANY-maze fear conditioning system), and after a 2-min adaptation period, the mice received three tone/foot shock pairings, where a tone (conditioned stimulus, 2.8 kH, 85 db, 30 s) always co-terminated with the foot shock (unconditioned stimulus, 2 s, 0.4 mA).

Techniques: Injection, Staining, Knock-Out, Immunoprecipitation, Western Blot, Control