Cortical sensory suppression during arousal is due to the activity-dependent depression of ...

13 Pages·2002·0.94 MB·English

Journal of Physiology(2002), 541.1, pp.319–331 DOI: 10.1113/jphysiol.2002.016857
© The Physiological Society 2002 www.jphysiol.org
Cortical sensory suppression during arousal is due to the
activity-dependent depression of thalamocortical synapses
Manuel A. Castro-Alamancos and Elizabeth Oldford
Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada H3A 2B4
The thalamus serves as a gate that regulates the flow of sensory inputs to the neocortex, and this gate
is controlled by neuromodulators from the brainstem reticular formation that are released during
arousal. Here we show in rats that sensory-evoked responses were suppressed in the neocortex by
activating the brainstem reticular formation and during natural arousal. Sensory suppression
occurred at the thalamocortical connection and was a consequence of the activity-dependent
depression of thalamocortical synapses caused by increased thalamocortical tonic firing during
arousal. Thalamocortical suppression may serve as a mechanism to focus sensory inputs to their
appropriate representations in neocortex, which is helpful for the spatial processing of sensory
information.
(Resubmitted 12 January 2002; accepted 13 February 2002)
Corresponding author M. A. Castro-Alamancos: Department of Neurology and Neurosurgery, Room WB210, Montreal
Neurological Institute, 3801 University Street, Montreal, Quebec, Canada H3A 2B4. Email: [email protected]
The thalamus serves as a gate that regulates the flow of VPM neurons send thalamocortical fibres to clusters of
sensory inputs to the neocortex, and this gate is controlled neurons located in layer IV (called ‘barrels’), and these
by neuromodulators from the brainstem reticular formation fibres also leave collaterals in upper layer VI (Jensen &
that are released during arousal (Steriade et al.1969, 1997; Killackey, 1987). Each barrel correlates on a one-to-one basis
Singer, 1977; Sherman & Koch, 1986; Castro-Alamancos, with the whiskers (Woolsey & Van der Loos, 1970). Despite
2002a,b). Among others, cholinergic and noradrenergic the anatomically modular and topographic arrangement,
fibres project to the thalamus (Hallanger et al.1990; Simpson the system displays extensive spatial and temporal
et al.1997). Neurons from these neuromodulatory systems integration. For instance, neurons in a given barrel column
discharge vigorously during behavioural arousal (Buzsaki yield the strongest response to a single principal whisker
et al.1988; Aston-Jones et al.1991), and the transmitters but also weaker responses to several surrounding whiskers
they release depolarize thalamocortical neurons and (Simons, 1978, 1985; Chapin, 1986; Armstrong-James &
enhance their firing rates (McCormick, 1992). Thus, during Fox, 1987; Moore & Nelson, 1998; Ghazanfar et al.2000;
aroused states of the brain, thalamocortical neurons display Petersen & Diamond, 2000). Inhibition in the neocortex
significantly enhanced spontaneous firing rates. Synapses has been implicated in the spatial contrast of principalvs.
are sensitive to activity and, in particular, thalamocortical adjacent whiskers (Simons, 1995). Also, the temporal
synapses display robust depression when stimulated at properties of neural responses in the barrel cortex have been
high rates (Castro-Alamancos, 1997; Gil et al.1997). These shown to modulate the size of the whisker representations
properties suggest that differences in the tonic firing rates (Sheth et al. 1998; Moore et al. 1999). In the rodent
of thalamocortical neurons between quiescent and aroused somatosensory system, receptive field and representation
states can change the gain of thalamocortical synapses and mapping have been carried out mainly in anaesthetized
significantly affect the mode of sensory transmission at the preparations where the level of arousal is similar to slow-
thalamocortical connection. wave sleep. However, during waking receptive fields and
pathways can change their properties at all levels of the
A useful model sensory system to investigate these issues is
sensory axis from the brainstem to the neocortex (Chapin
the rodent facial vibrissae (‘whisker’) system. Rats use their
& Woodward, 1981, 1982; Shin & Chapin, 1989, 1990;
whiskers to locate and identify objects (Guic-Robles et al.
Nicolelis et al.1993; Fanselow & Nicolelis, 1999).
1989; Carvell & Simons, 1990; Brecht et al.1997), and the
tactile skills of their whiskers are in some ways comparable The present study investigates how the primary thalamo-
to primates using their fingertips (Carvell & Simons, 1990; cortical pathway changes during aroused states. We show
Simons, 1995). The ventroposterior medial thalamus that sensory responses evoked in the barrel cortex by
(VPM) receives sensory information about the whiskers whisker stimulation are suppressed during aroused states.
from the trigeminal nucleus via lemniscal fibres (Chiaia et Sensory suppression in the barrel cortex is mainly a
al. 1991; Williams et al. 1994; Diamond, 1995). In turn, consequence of the activity-dependent depression of
320 M. A. Castro-Alamancos and E. Oldford J. Physiol.541.1
thalamocortical synapses caused by increased thalamo- Sensory stimulation
cortical tonic firing in VPM neurons during arousal. The sensory stimulation consisted of deflecting large caudal
Thalamocortical suppression during aroused states of the whiskers (one to four), which reliably discharged (>80% of trials
at 0.1Hz) the neurons recorded in VPM and barrel neocortex
brain may serve as a mechanism to focus sensory inputs to
with short latencies (3–7ms in VPM and 5–12ms in neocortex).
their appropriate representations (barrels) in neocortex,
The selected whiskers were inserted into a glass micropipette
which is helpful for the spatial processing of sensory
(1mm diameter) that was glued to the membrane of a miniature
information. speaker. Application of a 1ms square current pulse to the speaker
deflected the micropipette and the whiskers inside ~400mm.
METHODS Whisker stimulation was applied between 0.5 and 10s after the
reticular formation (RF) stimulation.
Surgical procedures
Current source–density analysis
Adult Sprague-Dawley rats (300g) were anaesthetized with
A 16-channel linear silicon probe (CNCT, University of
urethane (1.5gkg_1 I.P.) and placed in a stereotaxic frame.
Michigan, USA) was inserted into the barrel cortex perpendicular
Lidocaine (2%) was injected at incision sites and at points of
to the pial surface. This required insertion of the silicon probe at a
contact of the skin with the frame. A unilateral craniotomy
45deg angle (in the coronal plane) at 5.5–6mm lateral from the
extended over a large area of the parietal cortex. Small incisions
midline. Field potential recordings were obtained simultaneously
were made in the dura as necessary and the cortical surface was
from the 16 sites on the probe and from a VPM electrode that
covered with artificial cerebrospinal fluid (ACSF) containing (mM):
served to monitor multiunit activity. Band-pass filter settings were
NaCl 126; KCl 3; NaHPO 1.25; NaHCO 26; MgSO.7HO 1.3;
2 4 3 4 2 selected for field potential (1 Hz to 3kHz) or for multiunit
dextrose 10; CaCl.2HO 2.5. Body temperature was automatically
2 2 recordings (300Hz to 3kHz). A current source–density analysis
maintained constant with a heating pad. The level of anaesthesia
(CSD) was derived from the 16-channel cortical recordings, as
was monitored with field recordings and limb-withdrawal reflexes
previously described (Castro-Alamancos, 2000).
and kept constant at about stage III/3 using supplemental doses of
urethane (Friedberg et al.1999). At the end of the experiments the Chronic recordings
animals were killed with an overdose of sodium pentobarbitone Adult Sprague-Dawley rats (300g) were anaesthetized with sodium
(I.P.). The Animal Care Committee of McGill University, Canada, pentobarbitone (50mgkg_1I.P.) and placed in a stereotaxic frame.
approved protocols for all experiments. Lidocaine (2%) was injected at incision sites and at points of
contact of the skin with the frame. Recording electrodes were
Electrophysiological procedures
placed in the barrel cortex and stimulating electrodes were placed
Extracellular recordings were performed using electrodes
in the thalamic radiation. An insulated stainless-steel bipolar
(5–10MV) filled with ACSF; single units and field potentials were
recording electrode was placed in the whisker pad to record EMG
recorded simultaneously via the same electrodes located in the
signals. All electrodes and connectors were held in place using
VPM thalamus and the primary somatosensory neocortex (barrel
mini-screws and dental cement. During recovery from surgery the
cortex). When field potentials were recorded alone the electrode
animals were given Buprinorphine (0.02mgkg_1 S.C.). Animals
was placed at 800–1000mm from the surface. Field potential
were allowed 5_7days before testing and were recorded for
polarity is displayed as negative down. Coordinates (in mm, from
several days up to a maximum of 15days after surgery. During
bregma and the dura; Paxinos & Watson, 1982) for the VPM
recovery after surgery, animals were closely monitored for any
thalamus recording electrode were anterior–posterior=_3.5,
sign of distress or complications arising from the procedure.
lateral=3, depth=5–6. Coordinates (mm) for stimulating the
Electrophysiological recordings were performed as in anaesthetized
laterodorsal tegmentum (brainstem reticular formation; 100Hz,
animals, but JFET-operational amplifiers were attached to the
1s) were posterior=9, lateral=0.7, depth=5–6. The thalamic
recording electrodes at the animal’s head connector. During the
radiation was stimulated at approximately the following coordinates
recording sessions the animal was placed in an open field
(mm): posterior=3, lateral=4, depth=5. The medial lemniscus
containing photobeams that detected movements performed by
was stimulated at approximately the following coordinates
the animal. The field potential activity in the neocortex and the
(mm): posterior=5.5, lateral=1.5, depth=7.5. Electrical stimuli
motor activity detected with photobeams allowed us to differentiate
consisted of 200ms pulses of<200mA and were evoked using a
periods of active exploration from periods of slow-wave sleep. For
concentric stimulating electrode.
the population analysis, peak amplitudes of 20 randomly selected
Microdialysis thalamic radiation-evoked responses were measured per animal
To apply drugs into the neocortex during recordings a (n=10) and per condition (active vs. sleep). At the end of the
microdialysis probe (250mm diameter, 2mm long) was placed in experiments the animals were killed with an overdose of sodium
the neocortex 0.5–1mm medial from the recording electrode, as pentobarbitone.
previously described (Castro-Alamancos, 2000). ACSF was
continuously infused through the probe at 2–4mlmin_1. Drugs RESULTS
were prepared fresh, and protected from light and from oxidation
(40mM ascorbic acid in the ACSF) as required. Scopolamine, Thalamocortical suppression during activation
hexamethodide, phentolamine and propanolol were applied at Single-unit recordings were obtained simultaneously from
1–5mMeach, and CGP35348 (Novartis) was applied at 10mMin thalamocortical neurons of the VPM and from neurons in
ACSF . To apply TTX (2mMin ACSF) into the VPM thalamus a
layers III–IV of the primary somatosensory barrel neocortex
microdialysis probe (250mm diameter, 2mm long) was inserted
(Fig.1) of urethane-anaesthetized rats. Application of a train
at the following coordinates (mm): posterior=3, lateral=2–3,
depth=4–6. of electrical stimulation (100Hz, 1s) to the brainstem
J. Physiol.541.1 Thalamocortical suppression 321
Figure1. Activation induced by RF stimulation produces sensory suppression in neocortex
A, field potential (FP) and single-unit recordings obtained in the barrel cortex through the same electrode,
and a simultaneously recorded single unit in the VPM thalamus of a urethane-anaesthetized rat. RF
stimulation was delivered for 1s (100Hz) and produced a robust activating effect consisting of low
amplitude irregular activity in the cortical field potential, reduced firing in the cortical unit and enhanced
firing in the VPM unit. B, raw traces and binned sum data from 14 trials of sensory responses evoked by
whisker stimulation before (Control) and after RF stimulation. The cortical field and unit responses are
suppressed by RF stimulation, while the thalamic unit response is enhanced. C, cortical single-unit recording
obtained in the same experiment shown in Aand B. In contrast to cortical unit1 shown in Aand B, cortical
unit2 responds to RF stimulation by increasing its firing rate. However, like cortical unit1 this unit also
suppresses its response to whisker stimulation. Cortical unit2 was recorded after cortical unit1 in the same
penetration; the thalamic unit was the same for both cases.
322 M. A. Castro-Alamancos and E. Oldford J. Physiol.541.1
reticular formation (RF stimulation) produced a strong Thus, the probability of firing to whisker stimulation for
effect typical of aroused states (Fig. 1A) called activation, these short latency neurons was 88% during control
which is characterized by an electrographic sign of low conditions and 40% during activation induced by RF
amplitude fast activity (Moruzzi & Magoun, 1949). At the stimulation (48±3% reduction; P<0.0001, ttest; n=12
single-neuron level, during activation the firing rate of all units). The 48% reduction of very short latency cells is
VPM thalamocortical neurons recorded increased (n=55 significantly less than the 63% reduction observed for the
of 55; 100%), while the firing rate of the neocortical whole population of cells that includes cells with longer
neurons recorded either decreased (n=49 of 65; 75%) or latencies. In summary, during activation induced by RF
increased (n=16 of 65; 25%). VPM neurons increased stimulation the sensory response recorded in the barrel
their tonic firing after RF stimulation to 33±4Hz cortex is suppressed, while the sensory response recorded
(mean±S.D.) for several seconds. in the VPM thalamus is not suppressed.
In urethane-anaesthetized rats, whisker displacements using Suppression occurs at the thalamocortical
a mechanical stimulator produced successive sensory connection
responses in the VPM and barrel cortex (Fig. 1B). Single The barrel cortex is a complex structure that receives
units in VPM and in layer IV of barrel cortex responded afferents from the VPM thalamus in both layers IV and VI,
with short latency (3–7 and 5–12ms, respectively) and from where activity is distributed to other layers. To test
high fidelity (>80% probability of firing at short latency) which parts of this thalamocortical network are being
to whisker stimulation delivered at low frequencies (0.1Hz). suppressed by the RF stimulation we used a linear silicon
When brain activation was induced by RF stimulation, the probe containing 16 recording sites at 100mm intervals to
probability of firing to whisker stimulation at short latency record voltage throughout the layers of neocortex (Fig.2A)
(3–7ms) in the VPM increased from 82 to 100% (18±3% and derive a CSD in response to whisker stimulation
increase; P<0.0001, Student’s t test; n=55units). In (Bragin et al.2000; Castro-Alamancos, 2000). The current
contrast, in the barrel cortex the probability of firing to flow in the barrel cortex revealed by the CSD (Fig. 2B)
whisker stimulation at short latency (5–12ms) decreased showed that the sensory-evoked response corresponded to
from 82 to 19% (Figs 1B and 3A) (63±7% reduction; short latency current sinks in upper layer VI and layer IV,
P<0.0001, ttest; n=65units). Cortical single units that which spread horizontally within those layers and
enhanced or reduced their spontaneous firing in response vertically to layer III. Application of RF stimulation
to RF stimulation both showed a suppressed sensory- strongly depressed the sensory response in the barrel
evoked response during activation. Thus, cortical neurons cortex, but not the sensory response in the VPM (Fig.2B).
that enhanced their tonic firing as a consequence of RF Current flow in the neocortex was depressed beginning
stimulation depressed their response to whisker stimulation with the earliest (monosynaptic) sinks in layers VI and IV.
(Fig. 1C). The decrease in responsiveness to whisker As a consequence, the spread of activity within these layers
stimulation during activation was also reflected in the and to layer III was also strongly suppressed. On average
suppression of the sensory-evoked field potential response the peak amplitude of the short latency current sinks in
recorded in the cortex (57±8% reduction in amplitude; layers IV and VI and the longer latency sink in layer III
P<0.0001, t test; n=15; Figs 1B and 3B). The field were significantly depressed by 51.6±7, 59.8±7 and
potential response was recorded via the same electrode as 54.7±8%, respectively (n=3 experiments; P<0.0001, t
the single units and reflects the subthreshold synaptic test; Fig.3C). We also found that the response evoked in
activity of a population of neurons surrounding the the barrel cortex by stimulating thalamocortical fibres in
electrode. It is noteworthy that the single-unit responses the thalamic radiation was suppressed by RF stimulation
evoked by whisker stimulation showed on average a (Figs 2C and 3B; see below). In contrast, the response
stronger suppression than the field potential responses evoked in VPM by stimulating the primary sensory fibres
(Fig. 3A and B). This was a consequence of the fact that in the medial lemniscus was not suppressed by RF
some neurons such as the one in Fig. 1B almost entirely stimulation. The thalamic response evoked by medial
stopped responding to the sensory stimulus. It is likely that lemniscus stimulation has been characterized previously
some of these neurons still produced a subthreshold (Mishima, 1992). It consists of a very short latency and fast
response but we would not have been able to detect these component (arrow in Fig.2C) that is blocked by glutamate
responses using unit recordings. Other neurons may receptor antagonists (not shown) followed by a slower and
simply not respond at all after RF stimulation because they longer latency component (asterisk in Fig.2C) which is the
are only driven polysynaptically by the thalamic input and recurrent corticothalamic response, as demonstrated by
thus are entirely dependent on the firing of other cortical inactivating the neocortex (Mishima, 1992). RF stimulation
neurons, which may have been suppressed. This seems to did not significantly affect the initial fast response (Figs2C
be the case because cortical neurons responding with very and 3B; n=5 experiments; the peak amplitude of the medial
short latency (5–8ms) to whisker stimulation showed less lemniscus-evoked response was 1.1±0.08mV before and
suppression than the whole population of cortical neurons. 1.19±0.1mV after RF stimulation; not significant, ttest),
J. Physiol.541.1 Thalamocortical suppression 323
Figure2. Sensory suppression during activation occurs at the thalamocortical connection
A, schematic representation of the location of the 16-channel silicon probe placed at a 45deg angle in the
barrel cortex, which was used to record field potential responses through the layers of barrel neocortex. Also
note a single recording electrode placed in the VPM thalamus and a microdialysis probe located adjacent to
the recording electrode. The microdialysis probe was used to infuse TTX into the VPM as described in Fig.5.
B, current source–density analysis (CSD) of the sensory response evoked in the barrel cortex by whisker
stimulation before (Control) and after RF stimulation. The sink (red) and source (blue) distribution reveals
that the short latency responses in layers VI and IV are strongly depressed by RF stimulation. Also shown
below is multiunit activity from the VPM thalamus and a field potential recording from one of the cortical
sites (900mm in depth). The multiunit traces are the average of five sensory responses. Notice the depression
of the cortical response, but not of the thalamic response, after RF stimulation. The field potentials used to
derive the CSD are shown at the bottom. The scale range for the CSD is+3.5 to_3.5mVmm_2. C, overlaid
field potential responses showing the effect of RF stimulation (red traces) on cortical responses evoked by
whisker stimulation (left), cortical responses evoked by thalamic radiation stimulation (middle) and on
VPM responses evoked by medial lemniscus stimulation (right). The lemniscal response has two
components, marked by an arrow and an asterisk (see text for details). The responses are the average of ten
traces.
324 M. A. Castro-Alamancos and E. Oldford J. Physiol.541.1
but always abolished the long latency corticothalamic electrodes in the barrel cortex and thalamic radiation,
response that followed. Taken together, the results respectively. The animals were placed in an open field
indicate that sensory suppression is occurring at the (43cmw43cm) and motor activity was monitored using
interface between the thalamus and the neocortex, at the photobeams and an EMG electrode in the whisker pad.
thalamocortical connection. We found that indeed during behaviourally activated
states the thalamocortical response evoked by stimulating
Thalamocortical suppression occurs during
the thalamic radiation was suppressed. Figure 4 shows
behavioural arousal
recordings from a rat during two distinct behavioural
The experiments presented thus far were performed in
states: sleep and waking. During slow-wave sleep, as
anaesthetized animals, and activation was induced
indicated by the enhanced fast Fourier transform (FFT)
artificially by RF stimulation. Although RF stimulation
power of the spontaneous cortical activity at low frequencies
triggers wakefulness in sleeping animals (Lucas, 1975)
(<2Hz; Fig.4A), the thalamocortical-evoked response was
and mimics many of the features of the aroused brain
at its greatest level. As the animal awoke, the thalamo-
(Moruzzi & Magoun, 1949), the question remains whether
cortical-evoked response was strongly reduced, and was
suppression of the thalamocortical input actually occurs in
maintained at this reduced level during the vigourously
behaving animals during activated states. To test this
active period of exploration that followed. During waking
directly we chronically implantedrecording and stimulating
the FFT power showed an enhancement at 4–5Hz (Fig.4A).
This is probably theta activity picked up by volume
conduction from the cortical electrode because the FFT
analysis of the spontaneous activity did not distinguish
between negative and positive components. Sensory
suppression occurred when waking occurred spontaneously
or was triggered in a sleeping rat by the investigator. Based
on recordings from several behaving animals (n=10), the
amplitude of the field potential thalamocortical response
evoked by VPM stimulation was suppressed on average by
42±7% (P<0.0001, t test; n=10) between slow-wave
sleep and active exploration. In conclusion, similar to the
events that occur after RF stimulation, during behaviourally
activated states the thalamocortical response is suppressed
and therefore RF stimulation as used in the present study
mimics this aspect of natural arousal.
Mechanisms of thalamocortical suppression
How does thalamocortical sensory suppression induced
by RF stimulation occur? Thalamocortical synapses are
sensitive to activity and display pronounced activity-
dependent depression at frequencies above 1Hz (Castro-
Alamancos, 1997; Gil et al. 1997). Since RF stimulation
produces a strong activating effect in thalamocortical
neurons, which increases their firing rate, we reasoned that
increased thalamocortical activity caused by RF stimulation
Figure3. Population data showing the percentage could be depressing thalamocortical synapses and
changes induced by RF stimulation of VPM and cortex reducing the efficacy of the thalamocortical connection. If
responses
RF stimulation is depressing thalamocortical synapses by
A, percentage changes induced by RF stimulation of VPM and
increasing thalamocortical activity, then blocking thalamo-
cortex single-unit firing probability to whisker stimulation at short
cortical activity by inactivating the VPM thalamus should
latency intervals (3–7ms for VPM and 5–12ms for cortex). n=55
eliminate the suppressive effect of RF stimulation. VPM
and 65 units per group, respectively. *P<0.0001, ttest.
B,percentage changes induced by RF stimulation of field potential inactivation was produced with the sodium channel
responses evoked in cortex by whisker stimulation blocker tetrodotoxin (TTX), and was confirmed when
(WkråCortex) or thalamic radiation stimulation (TRåCortex) whisker-evoked responses were completely absent in the
and of responses evoked in VPM by medial lemniscus stimulation
neocortex (Fig.5A). To test the effect of RF stimulation on
(MLåVPM). n=15, 6 and 5 experiments per group, respectively.
the thalamocortical pathway before and after thalamic
*P<0.0001, ttest. C, percentage changes induced by RF
stimulation of current sink amplitudes evoked by whisker inactivation we stimulated the thalamic radiation. When
stimulation in layer IV, layer VI and layer III. n=3 experiments the thalamus was intact, the response evoked in the barrel
per group. *P<0.0001, ttest. cortex by stimulating the thalamic radiation was suppressed
J. Physiol.541.1 Thalamocortical suppression 325
by RF stimulation (Fig. 5B). However, when the VPM Thalamocortical synapses depress in response to activity
thalamus was inactivated with TTX, RF stimulation no and also in response to application of certain neuro-
longer suppressed the thalamic radiation-evoked response modulators in vitro(Gil et al.1997; Hsieh et al.2000) and
(Fig.5B). This indicates that sensory suppression induced by in vivo (Oldford et al. 2000). To distinguish between the
RF stimulation is a consequence of increased thalamo- two possibilities, an activity-dependent depression of
cortical firing in VPM. This experiment was performed thalamocortical synapses or a neuromodulator-mediated
several times (n=6 rats) with similar results. On average the depression of thalamocortical synapses, we tested whether
suppression of the thalamic radiation response was 55±7% TTX application in the VPM thalamus was affecting the
before (P<0.0001, ttest; n=6) and 6±4% after (P>0.1, cortical activating effects of RF stimulation. This was
ttest; n=6) TTX application, i.e. there was a 90% block of accomplished by comparing the power spectrums of
the effect of RF stimulation with thalamic inactivation. cortical activity in the presence and absence of thalamic
Figure4. Natural arousal produces thalamocortical suppression
A, fast Fourier transform (FFT) of the spontaneous field potential activity recorded from the barrel cortex of
a freely behaving rat. Blue indicates low power and red indicates high power for the frequency on the y-axis.
B,top: amplitude of the thalamocortical response evoked in the barrel cortex by stimulating the thalamic
radiation every 10s (open circles). The running averages of three successive responses are shown by filled
circles. Middle, amplitude of the electromyographic activity (EMG; arbitrary units) recorded from the
whisker pad with subcutaneous electrodes. Bottom, locomotor activity (arbitrary units) recorded by
photobeam detectors in the cage. The x-axis time scale corresponds to all graphs. The animal is sleeping for
the initial 11min (i.e. lying down in the cage with eyes closed) and the amplitude of the thalamocortical
response is large. After 11min, the rat wakes up and moves actively about the cage for the remainder of the
experiment, and the thalamocortical response is suppressed. C, traces correspond to a thalamocortical
response evoked during slow-wave sleep and during the active exploratory state that follows. Each trace
shown is 32.5ms. The arrows mark the onset of the electrical stimulus to the thalamic radiation.
326 M. A. Castro-Alamancos and E. Oldford J. Physiol.541.1
TTX (Fig. 5A). The results revealed that the activating synapses in the barrel neocortex, we applied simultaneously
effects of RF stimulation in the barrel cortex were not cholinergic (scopolamine and hexamethodide), nor-
significantly different before and during thalamic TTX adrenergic (phentolamine and propanolol) and GABA
B
application (n=6; ttest for the power between 0.5–15Hz, (CGP35348) receptor antagonists via a microdialysis
P>0.1). This would be expected if the activating effect of probe in the barrel neocortex. Application of this drug
RF stimulation in neocortex was mainly mediated by the combination in the cortex via microdialysis (Fig. 6)
basal forebrain (Jones, 1993). Since the modulation of significantly enhanced the amplitude of the whisker-
cortical neurons caused by RF stimulation was still present evoked response and made the response broader (1.4±
during VPM inactivation with TTX, we reasoned that 0.2mV beforevs. 2±0.3mV after the drug combination;
RF stimulation is not depressing the thalamocortical n=3 rats; P<0.0001, ttest). However, application of this
connection by releasing neuromodulator(s) in the cortex. drug combination did not block the sensory suppression
Thus, sensory suppression and cortical activation induced induced by RF stimulation (Fig. 6; n=3, suppression by
by RF stimulation are independent processes. Conversely, RF was 59±6% before and 75±5% after the drug
we propose that increased thalamocortical activity during combination).
activated states produces the depression of thalamo-
cortical synapses and consequently suppresses sensory- DISCUSSION
evoked responses in the neocortex. If this is the case,
The principal conclusion of the present study is that
activity in thalamocortical synapses should be able to
during aroused states the transmission efficacy of the
mimic the effect of RF stimulation. Indeed, similar to the
thalamocortical connection is reduced leading to the
effects of RF stimulation, repetitive stimulation at 10Hz
suppression of sensory responses in the neocortex. This is a
using sensory or thalamic radiation stimulation robustly
consequence of the activity-dependent depression of
suppressed thalamocortical responses (Castro-Alamancos
thalamocortical synapses caused by increased tonic firing
& Connors, 1996) to a similar extent as RF stimulation
of thalamic neurons. Importantly, this finding obtained
(Fig. 5C). To further test the potential for a cholinergic,
initially using brainstem RF stimulation was validated in
noradrenergic or GABAergic modulation of thalamocortical
Figure5. Sensory suppression induced by RF
stimulation is abolished by thalamic
inactivation
A, cortical field potential responses to whisker
stimulation (left traces) and to stimulation of the
thalamic radiation (right traces). The arrows mark
the onset of the whisker stimulus (left) and the
thalamic radiation electrical stimulus (right). The
numbers on the traces mark the locations on the plot
below. Infusion of TTX into the VPM thalamus
abolishes the cortical response to whisker
stimulation, but not the cortical response to
thalamic radiation stimulation. Also shown (right)
is a power-spectrum of the field potential activity
recorded in the cortex before (Control) and after RF
stimulation (RF stim) when the thalamus was intact
(continuous line) or inactivated with TTX (dashed
line). Thalamic inactivation does not significantly
affect the cortical activating effect of RF stimulation.
B,field potential responses to thalamic radiation
stimulation are suppressed by RF stimulation when
the thalamus is intact, but not when it is inactivated
with TTX. C, the thalamocortical response evoked
by stimulating the thalamic radiation is suppressed
by activity. Repetitive stimulation of the thalamic
radiation at 10Hz sharply depresses the
thalamocortical response (left), and this effect is
equivalent to RF stimulation in an intact thalamus
(right). The asterisk marks the small and long
latency response presumed to be due to intracortical
collaterals of corticothalamic cells (see Discussion
for details).
J. Physiol.541.1 Thalamocortical suppression 327
behaving animals. This indicates that the RF stimulation cortical pathway because this is what we found to be
used in the present study mimics the cortical sensory modified by RF stimulation. Accordingly, our behavioural
suppression that occurs during natural aroused states. experiments monitored only the thalamocortical sensory
pathway and not the sensory pathways to the thalamus or
As a sensory input travels upward from the periphery it is
brainstem. It is likely that additional modulatory systems,
not depressed by RF stimulation until it reaches the
which are activated during arousal or movement, produce
neocortex. In fact, at the level of the thalamus, sensory
further effects at the thalamic and brainstem levels. It
responses are enhanced by RF stimulation (Steriade et al.
seems also clear that the modulations that occur during the
1969; Singer, 1977; Castro-Alamancos, 2002a,b). CSD
waking state may be different depending on what the
revealed that the earliest current sinks in the thalamo-
animal is actually doing (Chapin & Woodward, 1981;
cortical recipient layers (IV and VI) of neocortex are
Fanselow & Nicolelis, 1999). Although this was not explored
suppressed by RF stimulation. The activity flow revealed
in detail in the present study, there are indications in Fig.4
by the CSD closely agrees with morphological studies,
that this is the case because the amount of thalamocortical
which have shown that thalamocortical fibres from VPM
suppression varied during arousal. The present study
project to layers IV–III leaving collaterals in layer VI of the
barrel neocortex (Bernardo & Woolsey, 1987; Jensen &
Killackey, 1987), and with electrophysiological studies
that mapped the laminar spread of whisker-evoked
activity within the neocortex using single-unit recordings
(Armstrong-James, 1995; Simons, 1995) and field potentials
(Di et al. 1990). The CSDs are also similar to those
obtained in primary somatosensory cortex using electrical
stimulation of the ventroposterior lateral thalamus (VPL)
(Castro-Alamancos & Connors, 1996; Kandel & Buzsaki,
1997). Since the thalamic output is enhanced and the earliest
current sinks in the thalamocortical recipient layers (IV
and VI) are suppressed, this indicates that sensory
suppression occurs at the thalamocortical connection.
This conclusion is supported by the observation that
cortical cells which enhanced or reduced their tonic firing
to RF stimulation both displayed a reduced sensory
response during arousal, indicating that a change in
cortical cell excitability cannot explain the suppression of
sensory responses. Taken together the results indicate that
cortical sensory suppression during arousal occurs at
thalamocortical synapses.
Previous work has shown that sensory responses are
reduced in the neocortex, thalamus and also brainstem
sensory nuclei during behaviourally aroused states and
movement in rodents, monkeys and humans (Chapin &
Figure6. Blocking cholinergic, noradrenergic and GABA
Woodward, 1981; Nelson, 1984; Cohen & Starr, 1987; Shin B
receptors in the neocortex does not abolish sensory
& Chapin, 1989, 1990; Fanselow & Nicolelis, 1999). These
suppression induced by RF stimulation
investigators have proposed that a central modulatory
A, field potential responses evoked in the neocortex by whisker
process must account for sensory suppression since it stimulation. Under control conditions RF stimulation suppresses
occurs away from the periphery and in the absence of the evoked response (upper traces). Simultaneous application of
actual motor activity. The present study shows that one scopolamine, hexamethodide, phentolamine, propanolol and
CGP35348 enhances the whisker-evoked response, but under these
mechanism which contributes to the suppression observed
conditions RF stimulation also suppresses the sensory-evoked
at the cortical level is the significantly increased thalamo-
response (lower traces). Traces are the average of five responses
cortical unit firing during arousal, which leads to activity-
from a representative experiment. B, population data from three
dependent depression of thalamocortical synapses. experiments in which the drugs mentioned in Awere applied. The
However, it is important to note that other factors that are average for each experiment was calculated from 10–15 control
not recruited by the RF stimulation used in the present traces and RF traces. RF significantly suppresses whisker-evoked
responses during control conditions and after application of the
study may also contribute to the changes previously
drug combination (*P<0.0001, ttest). Also, application of the
detected at the level of the thalamus and brainstem sensory
drugs significantly enhances the evoked response as compared to
nuclei. The present study focused only on the thalamo- control (**P<0.0001, ttest).
328 M. A. Castro-Alamancos and E. Oldford J. Physiol.541.1
emphasizes that during active behavioural states, such as thalamus is recurrently stimulated by corticothalamic
exploration, thalamocortical suppression is prevalent and synapses. However, this is not a problem because under these
that RF stimulation simulates this effect in anaesthetized same experimental conditions corticothalamic responses
animals. to low frequency stimulation are very small and only
corticothalamic stimulation above 5Hz produces a strong
The present study proposes that increased thalamocortical
thalamic response due to facilitation (Castro-Alamancos
unit firing produces activity-dependent depression of
& Calcagnotto, 2001). Moreover, the lack of involvement
thalamocortical synapses, which leads to sensory
of the corticothalamic connection in the cortical responses
suppression of neocortical responses. This conclusion is
evoked by thalamic radiation stimulation is demonstrated
based on several findings. First, thalamocortical synapses
by the fact that the amplitude of the cortical responses
depress with activity (Castro-Alamancos, 1997), and
to single stimuli of the thalamic radiation was not
neuronal tonic firing increases in thalamocortical neurons
significantly different before and after application of TTX
during arousal. After RF stimulation the firing rate of
in VPM (Fig. 5). (2) The other consequence of the
all thalamocortical neurons increased to ~33Hz. This
antidromic activation of layer VI corticothalamic neurons
discharge rate is characteristic of VPM neurons in awake
is that the cortex is recurrently stimulated via intracortical
behaving rats (Nicolelis et al.1993; Fanselow & Nicolelis,
collaterals from these neurons that reach the upper layers
1999). The analogous response of all VPM neurons to
(Zhang & Deschenes, 1997). Interestingly, these intracortical
RF stimulation was expected because thalamocortical
collaterals behave much the same way as corticothalamic
neurons represent a homogeneous population in their
synapses, producing strong facilitation (Stratford et al.
response to neuromodulators (McCormick & Prince,
1996) and long latency responses because these small
1987; McCormick, 1992). In addition, the differential effect
diameter fibres conduct much more slowly than the larger
of RF stimulation on the firing of neocortical neurons was
thalamocortical fibres (Ferster & Lindstrom, 1985; Swadlow,
also expected because in vitrostudies have shown distinct
1989). Consequently, we expected to observe a long latency
actions of neuromodulators depending on the cortical
response that could be attributed to these intracortical
neuronal type (McCormick & Prince, 1985; McCormick,
fibres when we stimulated at high frequencies. Indeed, as
1992; Xiang et al.1998).
shown in Fig. 5C (asterisk), what we found was a very
Second, blocking the firing of thalamocortical neurons in small, long latency response that followed the initial
the VPM with TTX is sufficient to eliminate the thalamo- thalamocortical response. The amplitude of this facilitated
cortical sensory suppression induced by RF stimulation by response to repetitive stimulation was about 5–10% of the
about 90%. A possible interpretation of this result is that it amplitude of the response we measured to single stimuli,
resulted from the block by TTX of the cortical activation and it was expected to be even smaller to single stimuli
mediated by the intralaminar nuclei of the thalamus because of the absence of facilitation. Thus, this leads to the
(Steriade et al. 1997) (e.g. by spread of TTX to the intra- conclusion that an intracortical component originating
laminar nuclei). However, this is unlikely for two reasons. from axon collaterals of corticothalamic cells would not be
(1) We found that cortical activation induced by RF present in the single stimuli responses we measured or it
stimulation is not different when the VPM thalamus is would be very small (<5% of the response) and have a
blocked with TTX. Thus, the cortical modulation induced long latency. Therefore, the response we measure in the
by RF stimulation is still present during application of cortex using single stimuli of the thalamic radiation is
TTX in VPM, although the thalamocortical suppression mostly (>90%), if not entirely, due to stimulation of
induced by RF stimulation is blocked. (2) It is unlikely that thalamocortical fibres.
the intralaminar nuclei were affected by the TTX because
Finally, neuromodulators that may be released in the
the distance between the microdialysis probe and the
neocortex by RF stimulation are known to affect
intralaminar nuclei is the same as that between the probe
thalamocortical synapses when applied in vivo(Oldford et
and the thalamic radiation. If TTX was spreading this
al. 2000) and in vitro (Gil et al. 1997; Hsieh et al. 2000).
distance the thalamic radiation-evoked responses should
However, we found that cholinergic, noradrenergic and
have been affected, which was not the case. Another
GABA receptor antagonists applied together in the
important consideration with this experiment is that due B
neocortex did not reduce sensory suppression induced by
to the need to inactivate VPM using TTX we had to use
RF stimulation (in fact, they slightly enhanced the
electrical stimulation of the thalamic radiation to stimulate
suppression), which demonstrates that these major neuro-
thalamocortical fibres. However, electrical stimulation of
transmitter systems do not contribute to thalamocortical
the thalamic radiation also evokes corticothalamic responses
sensory suppression induced by RF stimulation.
(Castro-Alamancos & Calcagnotto, 2001), which means that
layer VI corticothalamic neurons are being antidromically Taken together, the results of the present study lead to the
activated. There are two consequences of this. (1) The conclusion that increased thalamocortical activity in the