Research report
Ventrolateral periaqueductal gray lesion attenuates nociception but does not change anxiety-like indices or fear-induced antinociception in mice

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Abstract

The exposure of rodents to an open elevated plus-maze (oEPM: four open arms raised from the floor) elicits naloxone-insensitive antinociception. Midazolam infusion into the dorsal portion of the periaqueductal gray (dPAG), a structure of the descending inhibitory system of pain, failed to alter oEPM-induced antinociception. Chemical lesion of dorsomedial and dorsolateral PAG attenuated defensive behavior in the standard EPM (sEPM), an animal model of anxiety, but failed to change oEPM-induced antinociception. The present study investigated the effects of bilateral lesion, with the injection of NMDA (N-methyl-d-aspartic acid), of the ventrolateral column of PAG (vlPAG) (i) on nociceptive response induced by 2.5% formalin injected into the right hind paw (nociception test) in mice exposed to the enclosed EPM (eEPM: four enclosed arms – a non-aversive situation) or to the oEPM and (ii) on anxiety indices in mice exposed to the sEPM without prior formalin injection. Results showed that oEPM-induced antinociception was not altered by lesion of vlPAG. Nevertheless, the lesion reduced the nociceptive response in mice exposed to the eEPM and increased general locomotor activity during the eEPM and oEPM exposure. Furthermore, vlPAG lesion did not alter anxiety-like indices in mice exposed to the sEPM. The results suggest that vlPAG does not play a role in oEPM-induced antinociception or in defensive reactions assessed in the sEPM. Moreover, vlPAG inactivation induces pain inhibition in mice not exposed to an aversive situation and seems to increase general activity.

Research highlights

▶ Ventrolateral PAG lesion does not alter anxiety in mice. ▶ Ventrolateral PAG does not attenuate fear-induced antinociception. ▶ Ventrolateral PAG lesion attenuates pain in mice exposed to a non-aversive situation. ▶ Ventrolateral PAG plays a role in general activity in mice.

Introduction

Reynolds [1] was the first to report that electrical stimulation of the midbrain periaqueductal gray (PAG) produces pain inhibition. Since then, many studies have characterized the PAG as an important structure of the descending pain inhibitory system [for a review, see Ref. [2]]. The effect elicited by PAG stimulation on nociception involves modulation of the nociceptive transmission in the spinal cord, a role that seems to depend on the rostral ventromedial medulla (RVM) function, since RVM lesion abolishes this type of pain inhibition [2], [3], [4].

The PAG is anatomically and functionally organized, being divided into four longitudinal parts that run parallel to the aqueduct, viz. the dorsomedial, dorsolateral, lateral and ventrolateral columns [5], [6]. Chemical or electrical stimulation of dorsolateral and lateral PAG (dlPAG and lPAG) causes vigorous somatomotor activity, sympathoexcitation, cardio respiratory alterations and short-term non-opioid mediated analgesia [3], [5], [7], [8], [9], [10], [11]. On the other hand, the stimulation of the ventrolateral PAG (vlPAG) evokes “conservation-withdrawal” strategies or a passive coping reaction of quiescence/immobility, decreased vigilance and hyporeactivity, hypotension, bradycardia and an opioid analgesia which has a relatively long time course [5], [6], [7], [9], [10], [11], [12], [13].

It has been shown that PAG also participates in some types of environmentally induced analgesia [9], [14], [15], [16], [17]. Concerning the fact that animals display pain inhibition when confronted with threatening situations, Bolles and Fanselow [18] emphasized that this reaction has a clear adaptive value, since it gives the animal an opportunity to exhibit defensive behavior, even though an injury has occurred, thereby increasing its chances of survival. Thus, considered to be an important structure of the brain defensive system (e.g., Ref. [19], [20]), the PAG also participates in some types of aversive situation-induced antinociception.

It has been demonstrated that exposure of rodents to an elevated plus-maze (EPM: an apparatus in the shape of a plus sign, with two open and two enclosed arms, each with an open roof, elevated from the floor), a widely used animal model of anxiety [for a review, see Ref. [21]], besides inducing defensive behavioral responses, also elicits antinociception [22], [23], [24], [25]. In experiments carried out in our laboratory, EPM-exposed mice do not display intense antinociception, assessed by the formalin nociception test. Nevertheless, when placed in a totally open elevated plus-maze (oEPM: with four open arms) mice [26], [27] and rats [28] exhibit an intense antinociceptive response.

In order to investigate the role played by the dorsal portion of the PAG (dPAG: dm and dl columns) on oEPM-induced antinociception, we lesioned the dPAG by injecting a high dose of NMDA (N-methyl-d-aspartate) into this site. Bilateral dPAG lesion did not change oEPM-induced antinociception, but attenuated anxiety-like behavior in mice exposed to a standard EPM (sEPM: two open and two enclosed arms) [27]. Given that the vlPAG also modulates some forms of environmentally induced antinociception [9], [14], [16], [17], [29], the present study investigated the role of vlPAG on oEPM-induced antinociception and on sEPM-induced anxiety-like behavior in mice. To this end, a bilateral lesion was induced in the vlPAG by an injection of NMDA. The formalin test was used to assess nociception in the mice and was specifically chosen since it permits the assessment of pain sensitivity concurrently with exposure to an aversive situation, namely the oEPM.

Section snippets

Subjects

The subjects were male Swiss mice (São Paulo State University/UNESP, SP, Brazil), weighing 25–35 g. They were housed in groups of 7 per cage (41 cm × 34 cm × 16 cm) and maintained under a 12:12 h light/dark cycle (lights on at 7:00 a.m.) in a temperature (23 ± 1 °C) and humidity (55 ± 5%) controlled environment. Food and drinking water were freely available. All mice were experimentally naive, and experimental sessions were carried out during the light phase of the cycle (9 a.m.–5 p.m.).

Drugs

NMDA (N-methyl-d

Histology

All NMDA and sham lesions were verified histologically with Nissl staining. Similarly to previously described electrolytic [41] and chemical [27], [42] lesions of PAG, the lesion areas were extensive (see Figs. 1B and 2). In some cases, they included the lateral part of the PAG (lPAG) and also extended dorsally into the superior and/or inferior colliculi. The dorsal part of the PAG (dPAG, i.e. dorsolateral and dorsomedial columns), however, was left intact. There was no sign of lesion in the

Discussion

The results showed that oEPM-induced antinociception was not altered by bilateral lesion of vlPAG in mice. Surprisingly, vlPAG lesion decreased the nociception response in eEPM-exposed mice and increased locomotor activity in both eEPM and oEPM exposed-groups. Furthermore, in contrast to previously observed results with dPAG lesion [27], bilateral vlPAG lesion did not produce anxiolytic-like effects in sEPM-exposed animals that had not received a formalin injection.

Although the PAG has been

Acknowledgements

We thank Tatiani Sorregotti, Elisabete Zocal Paro Lepera, Rosana Finoti Pupim Silva and Cláudia Solano Rocha for the excellent technical assistance. This study was supported by FAPESP, CNPq and PADC/FCF-UNESP. J. Mendes-Gomes was supported by a FAPESP fellowship (05/01988-3), V.C.S. Amaral by a CNPq fellowship (142266/2008-6) and R.L. Nunes-de-Souza received a CNPq research fellowship (309407/2006-0).

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