Amygdalar neuronal activity mediates the cardiovascular responses evoked from the dorsolateral periaqueductal gray in conscious rats

doi:10.1016/j.neuroscience.2014.10.055

Neuroscience

Volume 284, 22 January 2015, Pages 737-750

https://doi.org/10.1016/j.neuroscience.2014.10.055 Get rights and content

Highlights

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Responses elicited by dlPAG activation require the neuronal activity in the BLA.
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Responses caused by the dlPAG activation depend on glutamatergic activity of BLA.
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The same role was not observed for the central amygdala.

Abstract

There is ample evidence that both lateral/dorsolateral periaqueductal gray (l/dlPAG) and basolateral amygdala (BLA) are essential for the regulation of the autonomic responses evoked during innate reactions to threatening stimuli. However, it is not well established to what extent the BLA regulates the upstream functional connection from the l/dlPAG. Here we evaluated the role of the BLA and its glutamatergic receptors in the cardiovascular responses induced by l/dlPAG stimulation in rats. We examined the influence of acute inhibition of the BLA, unilaterally, by injecting muscimol on the cardiovascular responses evoked by the injection of N-methyl D-aspartate (NMDA) into the l/dlPAG. We also evaluated the role of BLA ionotropic glutamate receptors in these responses by injecting antagonists of NMDA and AMPA/kainate receptor subtypes into the BLA. Our results show that the microinjection of NMDA in the BLA increased the mean arterial pressure (MAP) and heart rate (HR). Injection of NMDA into the l/dlPAG caused similar increases in these variables, which was prevented by the prior injection of muscimol, a GABA_A agonist, into the BLA. Moreover, injection of glutamatergic antagonists (2-amino-5-phosphonopentanoate (AP5) and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQ_X)) into the BLA reduced the increase in MAP and HR induced by l/dlPAG activation. Finally, the inhibition of the central amygdala neurons failed to reduce the cardiovascular changes induced by l/dlPAG activation. These results indicate that physiological responses elicited by l/dlPAG activation require the neuronal activity in the BLA. This ascending excitatory pathway from the l/dlPAG to the BLA might ensure the expression of the autonomic component of the defense reaction.

Introduction

It is generally agreed that specific neural circuits have developed in several organisms, that allow for a quick and strategic response to environmental threats. Specific pathways in the central nervous system (CNS) of mammals are activated during emotional or environmental stress, and induce various autonomic and endocrine responses (DiMicco et al., 2006, Szczepanska-Sadowska, 2008). These changes are accompanied by defensive behaviors including flight, freezing, defensive attack, and risk assessment (Blanchard and Blanchard, 1989a, Blanchard and Blanchard, 1989b), which favor survival.

One of the brain regions involved in the integration of specific physiological changes associated with emotional behavior is the periaqueductal gray matter (PAG) (Bandler et al., 2000). Studies have shown that chemical stimulation of glutamatergic neurons in the caudal portion of the lateral/dorsolateral periaqueductal gray (l/dlPAG) induces tachycardia and increases locomotor activity, blood pressure, and body temperature; responses which are similar to those produced during emotional stress (Carrive, 1993, Bandler and Shipley, 1994, de Menezes et al., 2009). Such tachycardic and pressor responses are mediated by efferent neuronal projections from the PAG to autonomic brain centers (Lovick, 1993, McNaughton and Corr, 2004).

Furthermore, the PAG has been defined as a lower center of defensive reactions influenced by downstream afferent projections arriving from the hypothalamus (Bandler and Keay, 1996, Bernard and Bandler, 1998). However, studies show that the PAG could also be a source of excitatory ascending projections to the hypothalamus during defensive reactions. In this regard, responses induced by stimulated l/dlPAG neurons are dependent on neuronal activity in the dorsomedial hypothalamus (DMH), suggesting that the PAG can produce changes in physiological parameters through upstream centers of neuronal activation, such as the hypothalamus (de Menezes et al., 2009, Horiuchi et al., 2009, Fontes et al., 2011).

Anatomical studies have also identified that both the dorsolateral (dlPAG) and the DMH form connections with the amygdala. Even though these connections appear to be indirect they play an important role in cardiovascular control during defense reactions. In fact, these three structures are believed to form an aversive system in the brain associated with anxiety (Graeff, 2004). The basolateral nucleus of the amygdala (BLA) has been identified as a site where GABAergic mechanisms play an important role in the regulation of cardiovascular function and behavior (Shekhar, 1993, Sanders and Shekhar, 1995). Additionally, it has been suggested that the BLA acts as an integration center for the anxiety states, and that glutamatergic neurotransmission is critical to the regulation of this condition (Campeau and Davis, 1995, Ledoux, 2012). Finally, glutamatergic stimulation of the BLA produces effects similar to those observed for the activation of l/dlPAG (Soltis et al., 1998). Altogether, these data suggest that the excitatory pathway between the dlPAG and the DMH could depend on the activation of BLA neurons.

There is ample evidence that the amygdala is critically involved in the regulation of innate and conditioned reactions to threatening stimuli (Ledoux, 1994, Campeau and Davis, 1995). Furthermore, it is frequently proposed that the PAG is downstream of the amygdala, directing motor outputs toward the proper defensive behavior (Ledoux, 2012). On the other hand, it is not well established to what extent the amygdala regulates the upstream functional connection from the l/dlPAG. Indeed, activation of the dlPAG leads to increased expression of c-Fos protein, a marker for neuronal activation, in the BLA (Ferreira-Netto et al., 2005).

Here, we evaluated role of the functional connection between the l/dlPAG and the BLA on the cardiovascular response evoked by the dlPAG. For that we examined the influence of acute inhibition of the BLA, by injecting muscimol, on responses evoked by the microinjection of the excitatory amino acid NMDA (N-methyl D-Aspartate) into l/dlPAG. We also evaluated the role of BLA ionotropic glutamate receptors in these responses by injecting 2-amino-5-phosphonopentanoate (AP5) and 6-cyano-7-nitroquinoxaline-2,3-dione) (CNQ_X), antagonists of NMDA and AMPA/kainate receptor subtypes, respectively, into the BLA. To establish the anatomical specificity of the effect of muscimol in the BLA, we also tested the consequence of identical injection into the central amygdala (CEA) in parallel experiments. Characterizing the interaction between PAG and amygdala in triggering the defense reaction will lead to a better understanding of these structures’ roles within a neural circuit that modulates reactions to threatening stimuli.

Section snippets

Ethical approval

All procedures were performed according to the regulations set forth by the National Institutes of Health Guidelines for the Care and Use of Laboratory Animals, and according to the journal policies and regulations on animal experimentation and were approved by the ethics committee for animal research of the Federal University of Ouro Preto (CEUA-UFOP; # 2010/26). All efforts were made to minimize the number of animals used in the present study, and to avoid any unnecessary distress to the

Stimulation of the BLA produces cardiovascular responses similar to those obtained by l/dlPAG activation

To confirm that the activation of BLA neurons is capable of producing increases in HR and MAP comparable to those induced by PAG activation, we injected an NMDA receptor agonist into the BLA. Injection of NMDA into the BLA produced increases in HR and MAP, relative to vehicle treatment (for treatment, HR: F_(1,96) = 16.89, p < 0.05; and MAP: F_(1,96) = 10.59, p < 0.05) (Fig. 1), as expected (Soltis et al., 1998). These changes were accompanied by increases in locomotor activity as previously reported (

Discussion

In this study, we evaluated the influence of BLA on the cardiovascular response produced by the activation of l/dlPAG. Our results showed that neuronal excitation in the l/dlPAG produces cardiovascular changes through the activation of glutamate receptors present in the BLA. Our data provide evidence that the BLA is downstream of the l/dlPAG, and directs the autonomic components of the defense reaction resulting from the activation of l/dlPAG.

First, we confirmed that the activation of glutamate

Conclusion

In this study, we showed that BLA activity plays a critical role in the modulation of physiological responses caused by l/dlPAG activation, and that these responses specifically depend on the glutamatergic activity of BLA neurons. Importantly, the same role was not observed for another region of the amygdala, the CEA. Thus, knowing that the autonomic responses evoked by l/dlPAG activation depends on neuronal activity in the DMH (de Menezes et al., 2009, Horiuchi et al., 2009), and that the

Acknowledgments

This work was supported by FAPEMIG, PRONEN, the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), and the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq).

References (49)

H.A. al Maskati et al.
Cardiovascular and motor components of the defence reaction elicited in rats by electrical and chemical stimulation in amygdala
J Auton Nerv Syst
(1989)
R. Bandler et al.
Columnar organization in the midbrain periaqueductal gray and the integration of emotional expression
Prog Brain Res
(1996)
R. Bandler et al.
Central circuits mediating patterned autonomic activity during active vs. passive emotional coping
Brain Res Bull
(2000)
R. Bandler et al.
Columnar organization in the midbrain periaqueductal gray: modules for emotional expression?
Trends Neurosci
(1994)
R.J. Blanchard et al.
Attack and defense in rodents as ethoexperimental models for the study of emotion
Prog Neuropsychopharmacol Biol Psychiatry
(1989)
P. Carrive
The periaqueductal gray and defensive behavior: functional representation and neuronal organization
Behav Brain Res
(1993)
L.G. da Silva et al.
Role of periaqueductal gray on the cardiovascular response evoked by disinhibition of the dorsomedial hypothalamus
Brain Res
(2003)
L.G.J. da Silva et al.
Excitatory amino acid receptors in the periaqueductal gray mediate the cardiovascular response evoked by activation of dorsomedial hypothalamic neurons
Neuroscience
(2006)
M.P. de la Mora et al.
Role of dopamine receptor mechanisms in the amygdaloid modulation of fear and anxiety: structural and functional analysis
Prog Neurobiol
(2010)
R.C. de Menezes et al.
Microinjection of muscimol into caudal periaqueductal gray lowers body temperature and attenuates increases in temperature and activity evoked from the dorsomedial hypothalamus
Brain Res
(2006)

A.R. de Oliveira et al.

Conditioned fear is modulated by D2 receptor pathway connecting the ventral tegmental area and basolateral amygdala

Neurobiol Learn Mem

(2011)

J.A. DiMicco et al.

Stress-induced cardiac stimulation and fever: common hypothalamic origins and brainstem mechanisms

Auton Neurosci

(2006)

C. Ferreira-Netto et al.

Neural segregation of Fos-protein distribution in the brain following freezing and escape behaviors induced by injections of either glutamate or NMDA into the dorsal periaqueductal gray of rats

Brain Res

(2005)

M.A. Fontes et al.

The dorsomedial hypothalamus and the central pathways involved in the cardiovascular response to emotional stress

Neuroscience

(2011)

P.L. Gabbott

Radial organisation of neurons and dendrites in human cortical areas 25, 32, and 32′

Brain Res

(2003)

P.L. Gabbott et al.

Areal and synaptic interconnectivity of prelimbic (area 32), infralimbic (area 25) and insular cortices in the rat

Brain Res

(2003)

C.G. Gentile et al.

The role of amygdaloid central nucleus in the retention of differential Pavlovian conditioning of bradycardia in rabbits

Behav Brain Res

(1986)

F.G. Graeff

Serotonin, the periaqueductal gray and panic

Neurosci Biobehav Rev

(2004)

T.S. Gray et al.

Peptide immunoreactive neurons in the amygdala and the bed nucleus of the stria terminalis project to the midbrain central gray in the rat

Peptides

(1992)

B.S. Kapp et al.

Amygdala central nucleus lesions: effects on heart rate conditioning in the rabbit

Physiol Behav

(1979)

J. Ledoux

Rethinking the emotional brain

Neuron

(2012)

J.E. Ledoux

The amygdala: contributions to fear and stress

Semin Neurosci

(1994)

T.A. Lovick

The periaqueductal gray-rostral medulla connection in the defence reaction: efferent pathways and descending control mechanisms

Behav Brain Res

(1993)

N. McNaughton et al.

A two-dimensional neuropsychology of defense: fear/anxiety and defensive distance

Neurosci Biobehav Rev

(2004)

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Amygdalar neuronal activity mediates the cardiovascular responses evoked from the dorsolateral periaqueductal gray in conscious rats

Highlights

Abstract

Introduction

Section snippets

Ethical approval

Stimulation of the BLA produces cardiovascular responses similar to those obtained by l/dlPAG activation

Discussion

Conclusion

Acknowledgments

J Auton Nerv Syst

Prog Brain Res

Brain Res Bull

Trends Neurosci

Prog Neuropsychopharmacol Biol Psychiatry

Behav Brain Res

Brain Res

Neuroscience

Prog Neurobiol

Brain Res

Neurobiol Learn Mem

Auton Neurosci

Brain Res

Neuroscience

Brain Res

Brain Res

Behav Brain Res

Neurosci Biobehav Rev

Peptides

Physiol Behav

Neuron

Semin Neurosci

Behav Brain Res

Neurosci Biobehav Rev