FIGURE LEGEND FOR SUPPLEMENTAL FIGURES
Figure S1. ND-evoked e-SP is associated with changes in the paired-pulse facilitation and can
be evoked by successive ND.
(A) Representative mean EPSCs (50 successive stimuli, including failures) evoked by paired-pulse
minimal stimulation before (basal) and after ND, and the scaled trace of the basal EPSC. (B) Pairedpulse facilitation (PPF) index before (basal) and after ND (n = 14). PPF index = [(2nd EPSC – 1st
EPSC) / 1st EPSC]). (C and D) Pr versus time from a representative synapse and averaged from 24
synapses, respectively. Arrows indicate the beginning of ND. (E) Relative changes from control
basal values of Pr before (basal) and after successive ND delivered every 9 minutes (n = 7 synapses
that displayed e-SP). *P < 0.05. Error bars indicate SEM.
Figure S2. Endocannabinoids released by pyramidal neurons elevate Ca2+ in adjacent
astrocytes.
(A) Pseudocolor images representing astrocyte fluorescence intensities before (basal) and after
evoking APs in the pyramidal neuron, which induced either e-SP or DSE (left and right). Scales bar,
20 μm. (B) Astrocyte Ca2+ spike probability evoked by APs that induced either e-SP (56 astrocytes
from n = 5 slices) or DSE (70 astrocytes from n = 6 slices) (left and right). Zero Time corresponds
to the beginning of AP train. (C) Astrocyte Ca2+ spike probability before (basal) and after APs in
control and AM251 (n = 5 and 6 slices for e-SP and DSE, respectively). (D) Astrocyte Ca2+ spike
probability evoked by ND in control and AM251 (37 astrocytes from n = 5 slices). Zero Time
corresponds to the beginning of the ND. (E) Astrocyte Ca2+ spike probability (left) and Ca2+
oscillation frequency (right) before (basal) and after ND in control and AM251 (n = 5 slices). (F)
Proportion of astrocytes that increased their intracellular Ca2+ by ND in control and AM251 (37
astrocytes from n = 5 slices). * P < 0.05 and ** P < 0.01. Error bars indicate SEM.
Figure S3. ND-evoked e-SP occurred at 24 and 34 ºC.
(A) Synaptic parameters at 24 and 34 ºC before (basal; open circles) and after ND (filled circles) (n
= 6 synapses). Red symbols represent mean values. (B) Relative changes from basal values at 24 ºC
of synaptic parameters before (basal) and after ND at 24 and 34 ºC (n = 6). *P < 0.05 and **P <
0.01. Error bars indicate SEM.
Figure S4. Endocannabinoid-induced DSE is unaffected by thapsigargin.
(A) Pr versus time in control and after superfusion with 1 μM thapsigargin (n = 5 synapses). Zero
time corresponds to the beginning of ND. (B) Relative changes from control basal values of
synapticparameters before (basal) and after ND in control and thapsigargin (n = 5). *P < 0.05. Error
bars indicate SEM.
Figure S5. Astrocyte Ca2+ elevations increase transmitter release through a CB1R-insensitive
and mGluR-dependent mechanism.
(A) Schematic drawing depicting paired recordings from one pyramidal neuron and one astrocyte,
and the electrode for synaptic minimal stimulation. (B) Pseudocolor images representing
fluorescence intensities of a fluo 4-filled astrocyte loaded with the Ca2+-cage NP-EGTA before
(basal) and after UV-flash photolysis. Scale bar, 20 µm. (B) Pr versus time in control, AM251 (n =
8), and MPEP+LY (n = 5). Zero time corresponds to the UV-flash photolysis of NP-EGTA. (C)
Relative changes from control basal values of synaptic parameters before (basal) and after astrocyte
Ca2+ elevations by UV-flash photolysis of NP-EGTA in control, AM251 (n = 8), and MPEP+LY (n
= 5). *P < 0.05, **P < 0.01. Error bars indicate SEM.
Figure S6. mGluR activation modulates synaptic transmission.
(A) Pr versus time after a fast puff application of DHPG by a 0.5 s pressure pulse (arrows), after
slow bath perfusion of the slice with DHPG, and after washout. (B) Relative changes of synaptic
parameters from control basal values after puff application of DHPG, bath perfusion with DHPG,
and after washout (n = 5). (C) Responses evoked by minimal stimulation (20 consecutive stimuli;
top traces) and averaged EPSCs (n = 50 stimuli; bottom traces) before (basal) and after puff
application of DHPG in control and in the presence of the group I mGluR antagonists MPEP +
LY367385. (D) EPSC amplitudes from a representative synapse in control and MPEP + LY367385.
(E) Pr versus time after a fast puff application of DHPG (arrows) in control and after perfusion with
MPEP + LY367385 (n = 5). (F) Relative changes from control basal values of synaptic parameters
before (basal) and after puff application of DHPG in control and in the presence of MPEP +
LY367385 (n = 6). *P < 0.05, **P < 0.01. Error bars indicate SEM.
Figure S7. Time course of the ND-evoked DSE in homoneuronal and heteroneuronal
synapses.
(A) Synaptic efficacy, Pr and synaptic potency versus time in homoneuronal synapses (green
circles; n = 14) and heteroneuronal synapses (blue circles; n = 14) that showed DSE upon ND. Zero
time corresponds to the beginning of ND.