10.5061/DRYAD.RN8PK0P7K
Bowen, Anna
0000-0002-8911-2572
University of Washington
Chen, Jane
University of Washington
Huang, Y. Waterlily
0000-0002-9147-8418
University of Washington
Baertsch, Nathan
0000-0003-1589-5575
Seattle Children's Research Hospital
Park, Sekun
0000-0003-0634-590X
University of Washington
Palmiter, Richard
University of Washington
Dissociable control of unconditioned responses and associative fear
learning by parabrachial CGRP neurons
Dryad
dataset
2020
National Cancer Institute
https://ror.org/040gcmg81
R01-DA24908
National Institute of General Medical Sciences
https://ror.org/04q48ey07
T32NS099578
2020-08-28T00:00:00Z
2020-08-28T00:00:00Z
en
709185 bytes
2
CC0 1.0 Universal (CC0 1.0) Public Domain Dedication
Parabrachial CGRP neurons receive diverse threat-related signals and
contribute to multiple phases of adaptive threat responses in mice, with
their inactivation attenuating both unconditioned behavioral responses to
somatic pain and fear-memory formation. Because CGRPPBN neurons respond
broadly to multi-modal threats, it remains unknown how these distinct
adaptive processes are individually engaged. We show that while three
partially separable subsets of CGRPPBN neurons broadly collateralize to
their respective downstream partners, individual projections accomplish
distinct functions: hypothalamic and extended amygdalar projections elicit
assorted unconditioned threat responses including autonomic arousal,
anxiety, and freezing behavior, while thalamic and basal forebrain
projections generate freezing behavior and, unexpectedly, contribute to
associative fear learning. Moreover, the unconditioned responses generated
by individual projections are complementary, with simultaneous activation
of multiple sites driving profound freezing behavior and bradycardia that
are not elicited by any individual projection. This semi-parallel,
scalable connectivity schema likely contributes to flexible control of
threat responses in unpredictable environments.
Data were collected for behavioral measures, intersectional anterograde
tracing, slice electrophysiology, and autonomic measures. Data for
behavioral measures were collected using video and have been fully
processed: freezing behavior was measured based on absence of locomotion
and is given as a semi-continuous measure across session epochs. Anxiety
measures were obtained by tracked video data comparing time spent in
various locations within an elevated plus maze. Excel sheets summarizing
the various measures across individual experimental and control animals
have been provided. Intersectional anterograde tracing data were collected
from sectioned brain tissue from mice injected with retrogradely
transported Flp in distinct target regions, and INTRSECT viruses
restricting expression to Cre-expressing neurons either expressing or not
expressing Flp injected into the PBN. All regions of interest were
sectioned such that each imaged section was 180um apart and were mounted
and imaged in caudal to rostral order. Each section that contained an area
of interest was imaged and analyzed for each experimental animal. Data
were processed by outlining each downstream target region with an ROI in
imageJ with an off-target ROI from the same section for
background-subtraction. Mean pixel intensity was measured within the ROI,
then this value had the background intensity subtracted before being
multiplied by ROI size to generate a value for total pixel intensity for
that ROI. Both the mean pixel intensity, backgroun intensity, ROI size,
and total pixel intensity are included for each section of each
target-region. Slice electrophysiology data consists of the mean EPSC
frequencies and amplitudes obtained from current clamp recordings while
delivering light pulses at terminals expressing excitatory or inhibitory
opsins and recording from putative post-synaptic neurons. Autonomic data
was collected using pulse oximeters with collar sensors or in a
plethysmography chamber for some respiratory measures in awake, freely
moving animals. Reported data is the average heart rate, blood
oxygenation, respiratory rate, and pulse distension during baseline and
stimulation periods. Vasodilation measures are the result of analyzing
infrared camera images of mice where ROIs were drawn to extract
temperature readings 1/3 below base of tail and measuring temperature
difference before and during 30-Hz photostimulation. Original infrared
image files are also provided.
Columns/rows of data for excluded animals have been highlighted in yellow.