Brain Research 880 2000 1–10 www.elsevier.com locate bres
Research report
Expression and conditioned inhibition of fear-potentiated startle after stimulation and blockade of AMPA Kainate and GABA receptors in
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the dorsal periaqueductal gray
Markus Fendt
¨ ¨
¨ Tierphysiologie
, Universitat Tubingen, Auf der Morgenstelle 28, D-72076 Tubingen, Germany Accepted 20 June 2000
Abstract
Previous work showed that the dorsal periaqueductal gray is involved in the inhibition of fear-potentiated startle. The present study investigated the effects of blockade and stimulation of Kainate AMPA and GABA receptors within the dorsal periaqueductal gray on
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expression and conditioned inhibition of fear-potentiated startle. Blockade of the Kainate AMPA receptors enhanced whereas stimulation of the Kainate AMPA receptors decreased expression of fear-potentiated startle. These effects do not reflect conditioned inhibition since
this modulation was not changed by injections of Kainate AMPA receptor agonists or antagonists into the dorsal periaqueductal gray. Stimulation and blockade of GABA receptors within the dorsal periaqueductal gray neither affected expression of fear-potentiated startle
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nor its conditioned inhibition. The present results together with findings from the literature indicate that glutamate in the dorsal periaqueductal gray is a critical substrate for the expression and modulation of fear-related behaviours.
2000 Elsevier Science B.V. All
rights reserved.
Theme : Neural basis of behavior
Topic : Motivation and emotion
Keywords : Acoustic startle response; Anxiety; Conditioned fear; Conditioned inhibition; GABA; Glutamate; Periaqueductal gray
1. Introduction 18,22,42,43]. One possibility to inhibit fear-potentiated
startle is conditioned inhibition which was established by The inhibition of fear is important in situations where it
Falls and co-workers [16,19]. In their procedure, the fear would lead to inappropriate behaviour, e.g. when an
conditioning training pairings of a light CS with a organism recognises that a threat is over. The knowledge
footshock US is followed by a further training in which of the neurochemical and neuroanatomical basis of this
the rats learn that a combination of a conditioned inhibitor mechanism might help to develop strategies to treat
CI: e.g. a noise with the CS is not associated with an US, pathological fear in humans. One of the most valuable
but the CS alone further predicts a US. After these training models to investigate the neural basis of fear in humans
procedures, the ASR amplitude is still potentiated in the and
rats is
the fear-potentiated
startle paradigm
presence of the CS alone, whereas the combination of the [8,12,24,32]. Here, the acoustic startle response ASR is
CI and the CS inhibits fear-potentiated startle. The CI potentiated in the presence of a conditioned stimulus CS:
alone does not affect the ASR amplitude. e.g. a light which has previously been paired with an
A series of studies were carried out to investigate the aversive, unconditioned stimulus US: e.g. a footshock.
neural basis of conditioned inhibition of fear-potentiated During the last years, the investigation of inhibition of
startle. A c-fos study [9] pointed to several brain regions fear
has received
increasing interest
e.g., [9,16–
possibly being involved in the inhibition of fear. Most of these and other regions were lesioned during the last years
without effects on conditioned inhibition of fear-poten-
Tel.: 149-7071-2975347; fax: 149-7071-292618. E-mail address
: markus.fendtuni-tuebingen.de M. Fendt.
tiated startle e.g., [17,18,23,26]. Two recent studies gave
0006-8993 00 – see front matter
2000 Elsevier Science B.V. All rights reserved. P I I : S 0 0 0 6 - 8 9 9 3 0 0 0 2 6 6 5 - 2
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first hints to brain structures possibly involved in con- available. The experiments were done in accordance with
ditioned inhibition of fear-potentiated startle: 1 Fendt ethical guidelines for the care and use of experimental
[22] showed that a blockade of GABA receptors within
animals and were approved by the local council of animal
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¨ ¨
the dorsal region of the periaqueductal gray dPAG care Regierungsprasidium Tubingen, ZP 4 96.
attenuates expression of conditioned inhibition of fear- potentiated startle; 2 Heldt and Falls [29] demonstrated
2.2. Surgery that electrolytic lesions of the brachium of the colliculus
inferior block the expression of conditioned inhibition of The animals were anaesthetised with ketamine xylazine
fear-potentiated startle. 9:1, 100 mg kg, i.p.. Two stainless guide cannulae
Whereas the brachium of the colliculus inferior is a part diameter: 0.7 mm were implanted bilaterally into the
of the auditory pathway mediating the noise CI [37,50], the brain using following stereotaxic coordinates of Paxinos
dPAG is a brain structure which is not directly involved in and Watson [39]: rostrocaudal: 26.3 mm, mediolateral:
the processing of the auditory CI, but is well known for 6
0.6 mm, dorsoventral: 25.0 mm relative to Bregma. modulation of fear and anxiety [5,24,48]. Beside its
The guide cannulae were fixed to the skull with dental possible role in conditioned inhibition of fear-potentiated
cement and three anchoring screws. After the surgery and startle [22], the dPAG plays a role in the expression of
between the experiments, stainless steel stylets diameter: fear-potentiated startle. Walker and colleagues [49] demon-
0.4 mm were inserted into the guide cannulae to maintain strated that dPAG stimulation by injections of Kainic acid
patency. All subjects were allowed 4 days recovery before block fear-potentiated startle whereas dPAG lesions pre-
training and testing. vent the loss of fear-potentiated startle seen after training
with high intensity USs [11]. Furthermore, we previously 2.3. Apparatus
demonstrated [25] a projection from the dPAG to the caudal pontine reticular nucleus, a crucial part of the
The rats were trained in two identical, acoustically
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primary startle circuit [32]. A number of other studies isolated dark boxes 38360328 cm . The floor of the
using different animal models to investigate fear-related boxes was composed of stainless steel bars spaced |15
behaviours showed that the dPAG is strongly involved in mm apart, through which the unconditioned stimulus US,
the expression and modulation of fear [20,28,30,31,34– a 0.5 s, 0.6 mA footshock could be administered. The
36,44]. It is important to note that with the methods used footshocks were produced by a shock generator custom-
¨ in the studies cited above it is not possible to distinguish
made at the University of Tubingen located outside the between the dorsomedial and the dorsolateral part of the
chamber. The conditioned stimulus CS for fear-con- PAG, two areas which can be clearly differentiated [2].
ditioning paired with the US was a 3.7 s white light
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The aim of the present study was to investigate the role produced by a 15 W bulb light
trials, the conditioned of AMPA Kainate and GABA receptors within the dPAG
inhibitor CI, predicting that the light is not followed by a
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in the expression and conditioned inhibition of fear-poten- footshock, was a 3.7 s, 70 dB white noise produced by a
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tiated startle. First, we tested whether the blockade of loudspeaker noise
→ light
trials. The bulbs and the fear-potentiated startle which were reported after injections
loudspeakers were located on the top of the boxes. The of the AMPA Kainate receptor agonist Kainic acid into the
presentation of the US, CS and CI were controlled by a dPAG reflects conditioned inhibition. In addition, the
microcomputer and an appropriate interface Hortmann effects of injections of the AMPA Kainate receptor an-
universal function synthesiser, Hortmann, Neckartenzling- tagonist NBQX were investigated. Second, we attempted
en, Germany. to replicate the blockade of conditioned inhibition of fear-
To measure fear-potentiated startle and conditioned potentiated startle reported after GABA receptor blockade
inhibition, the animals were tested in two identical test
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by Picrotoxin within the dPAG and to confirm these results chambers. The rats were placed in wire mesh cages 203
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by an opposite effect after GABA receptor stimulation by 10312 cm with a steel floor, which were put up on a
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Piperidine. piezoelectric accelerometer custom-made at the University
¨ of Tubingen. The accelerometer was located inside a
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sound-attenuated test chamber 100380360 cm . Move-
2. Materials and methods ments of the rats resulted in changes of the voltage output
