SRI Supplement to Reproductive Sciences - Volume 25 Number 1 - March 2018 - 191A

Scientific Abstracts

F-026

F-027
Term Contractions Characterized by an Abrupt Increase in Uterine
Bioelectrical Activity. Roger Young*,1 Lauren A Miller†,2 Courtney
Olsen-Chen,2 Eva Pressman,2 Neil Seligman.2 1PreTel Inc., Chattanooga,
TN, United States; 2University of Rochester, Rochester, NY, United States.
INTRODUCTION: The uterine bioelectrical changes preceding labor
have not previously been described. In this study the bioelectrical activity
of the human uterus was observed to determine if the onset of labor is
preceded by a gradual increase in uterine bioelectrical activity.
METHODS: A single subject (no prior preterm birth) underwent 2x
weekly uterine monitoring from 35-39 weeks using multichannel EMG,
with directional, or "area", EMG sensors (previously shown to provide
enhanced signal acquisition vs. pad-type sensors). Uterine activity was
recorded for 30 mins/session, with concomitant tocodynamometry (toco).
Three classes of electrical activity were analyzed: electrical "spikes"(>50
µV for 3-15sec); "brief bursts"(≥ 3 spikes in ≤20 sec); "prolonged bursts"
(a single spike lasting >15 sec, or bursts lasting >20 sec). Correlations
between electrical activity classes and both advancing gestational age
(GA) and contractions frequencies were computed using Spearman's
correlation coefficients.
RESULTS: The patient underwent 10 sessions (details table 1). Session 9
and 10 were recorded during labor. A correlation between prolonged bursts
and both spikes (r2=0.7, P=0.04) and GA (r2=0.7, P=0.03) were noted.
As seen in Image 1-2, the curve is exponential, with a high frequency
of prolonged bursts being first observed at 39 weeks. A high correlation
between the frequency of contractions on toco and the frequency of
prolonged bursts was seen (r2=0.44), but was not significant (P=0.2).

191A

CONCLUSION: Prolonged bursts were absent and spikes were
infrequent before 39 weeks, even when the subject experienced
contractions. Concomitant with the onset of labor, there is an abrupt
increase in the frequency of prolonged bursts and spikes, not a gradual
week-by-week increase. Contractions at earlier gestations were seen on
toco, but the electrical activity is not equivalent to that seen at 39 weeks.
This suggests the electrical signaling behind labor contractions and
"pre-labor" or "Braxton-Hicks" contractions are fundamentally different.
*Figure(s) will be available online.

F-028
A Novel Method Using Fluorescence Resonance Energy Transfer
(FRET) Imaging and Targeted FRET Reporters to Investigate
cAMP Function in Human Myometrial Cells. Alice Varley†,1 Andreas
Koschinski,2 Mark Johnson,1 Manuela Zaccolo*.2 1Imperial College
London, London, United Kingdom; 2University of Oxford, Oxford, United
Kingdom.
INTRODUCTION: cAMP regulates multiple complex signalling
networks. Subcellular compartmentalisation of cAMP has emerged as
the mechanism determining its ability to activate numerous hormonespecific cellular functions. Tight regulation of the spatial and temporal
dynamics of cAMP signalling is understood to determine specificity. FRET
imaging is a powerful tool used to study cAMP dynamics in living cells.
FRET biosensors monitor cAMP concentrations and activity of cellular
effectors, such as PKA, which mediate compartmentalised signalling
cascades. Targeted sensors measure changes in the cAMP signal at
explicit subcellular locations. Currently, there is limited data visualising
the cAMP signalling events in human myometrial cells. Our aim is to
determine whether cAMP action is confined to distinct microdomains
within myometrial cells using targeted FRET reporters and the functional
significance of this with the onset of labour.
METHODS: Human primary myometrial cells from term no labour
women undergoing caesarean section (n=3) were infected with adenoviral
vectors encoding for either a cytosolic sensor (EPAC-SH187) or a version
that is targeted to the plasma membrane via fusion to the scaffold protein
AKAP79 (AKAP79-CUTie). Cells expressing the sensors were excited
at a wavelength of 430nm and a stimulus applied. The subsequent ratio
signal was monitored using OptoFluor software measuring the mean
intensity values of the resulting fluorophore emissions of the cyan and
yellow fluorescent proteins. Data was statistically analysed following
background correction to calculate the ratio between the fluorophore
mean intensities. These ratio values correlate to changes in intracellular
cAMP concentration.
RESULTS: Initial optimisation studies demonstrated successful infection
with differing concentrations of the H187 virus. The mean FRET response
expressed as a percentage change from the baseline show that maximum
FRET change in the primary myometrial cells using the EPAC-SH187
sensor following agonist stimulation was 263%. This result is comparable
to similar findings using neonatal cardiomyocytes. Further optimisation
studies are needed to attain a higher infection efficiency using the targeted
FRET sensor, AKAP79-CUTie.
CONCLUSION: Although extensive research has focused on the
molecular mechanisms and pathways co-ordinating cAMP activity,
multiple aspects of its functionality need further investigation, particularly
in the myometrium. Advancements in FRET imaging techniques have
enhanced understanding of cAMP signalling events, principally in
cardiomyocytes. Our aim is to enhance understanding of cAMP's role in
the myometrium, specifically whether its action is limited to microdomains
within the cell and the functional significance of these domains.

Friday Posters

Maternal Oxytocin Administration at Term Affects Uterine Gap
Junction-Associated Gene Expression in Mice. Shelly Soni†,2 Prodyot
K Chatterjee,1 Frances F Hsieh,2 Xiangying Xue,1 Burton Rochelson*,2
Christine N Metz*.1,2 1The Feinstein Institute for Medical Research,
Manhasset, NY, United States; 2Zucker School of Medicine at Hofstra/
Northwell, Manhasset, NY, United States.
INTRODUCTION: Oxytocin is a neuropeptide/hormone associated with
parturition. Exogenously administered oxytocin is commonly used to
induce and augment labor. We investigated the effects of maternal oxytocin
administration on uterine gap junction-associated gene expression in
mice at term.
METHODS: Pregnant C57BL/6 mice (10 weeks old, gestational
day 18) were given either saline (100μl) or OXT (1U/mouse, 100μl)
subcutaneously every 30min for 2hrs (4 per group); 6hrs after the 1st
injection dams were euthanized and pups were delivered by Cesarean
section. Uterine specimens from the 2 horns closest to the cornual end
were collected, snap frozen in liquid N2, and stored at -80°C. RNA was
extracted and quantified using the NanoDrop. Uterine RNA samples
were converted to ds-cDNA and gene expression was analyzed by qPCR
with the RT2 Profiler for mouse gap junctions (Qiagen) using the Roche
LightCycler 480, according to the manufacturer's instructions.
RESULTS: We identified 3 genes that were significantly up-regulated
and 4 genes that were significantly down-regulated in term uteruses
following maternal oxytocin administration (vs. saline) (p< 0.05). Gja6
(connexin 33) (p< 0.03), Gnai1 (G protein, alpha inhibiting 1)(<0.05)
and Prkg2 (protein kinase, cGMP-dependent, type II)(p<0.001), which
are involved in gap junction signaling, were significantly down-regulated
by oxytocin. In addition Plcb1 (phospholipase C, beta 1) (p<0.05), which
plays a role in oxytocin-mediated uterine contractility and Pdgfra (plateletderived growth factor receptor, alpha)(p<0.001), which is involved in
prostaglandin synthesis, were significantly up-regulated by oxytocin.
CONCLUSION: Exogenously administered oxytocin just prior to
labor in the mouse results in the differential regulation of gap junctionassociated genes in the uterus. Gap junction-associated genes involved
in prostaglandin synthesis, as well as oxytocin contractility pathways
are up-regulated, while genes involved in smooth muscle relaxation (e.g.
Prkg2 and Gnai1) are down-regulated. These opposing effects should
promote synchronized uterine contractility.
*Figure(s) will be available online.

Reproductive Sciences Vol. 25, Supplement 1, March 2018

Table of Contents for the Digital Edition of SRI Supplement to Reproductive Sciences - Volume 25 Number 1 - March 2018