Intra-visceral insular cortex 2-arachidonoylglycerol, but not N-arachidonoylethanolamide, suppresses acute nausea-induced conditioned gaping in rats
Introduction
Unlike emetic animal models, rodent models of nausea have remained rather elusive. Traditional consumption tests following flavor-illness associations, previously thought to reflect nausea, have revealed that rats avoid a distinct taste that has been paired not only with emetic agents, but also rewarding drugs (Parker, 1995). On the other hand, conditioned gaping, as assessed using the taste reactivity (TR; Grill and Norgren, 1978a) test, is a more reliable model of nausea as it is selectively produced by emetic drugs (Parker, 2003; also see Parker, 2014). Moreover, classic anti-emetic treatments such as the 5-hydroxytryptamine (5-HT) 3 receptor antagonist, ondansetron (Limebeer and Parker, 2000), have been shown to prevent the establishment of conditioned gaping (without interfering with taste avoidance), as do synthetic and plant-based cannabinoid agonists (Parker and Mechoulam, 2003). More recently, enhancement of the endogenous cannabinoids (eCBs), N-arachidonoylethanolamide (anandamide; AEA; Devane et al., 1992) and 2-arachidonoylglycerol (2-AG; Mechoulam et al., 1995, Sugiura et al., 1995) by blockade of their respective catabolic enzymes, fatty acid amide hydrolase (FAAH; Cravatt et al., 1996), and monoacylglycerol lipase (MAGL; Dinh et al., 2002) have been shown to protect against nausea (Parker et al., 2011, Sharkey et al., 2014). Specifically, the FAAH inhibitor, URB597, was found to suppress acute nausea-induced conditioned gaping (Cross-Mellor et al., 2007), as did systemic administration of the novel MAGL inhibitor, MJN110 (Parker et al., 2014), or pretreatment with exogenous 2-AG alone (Sticht et al., 2012).
Although the emetic circuitry mediating the vomiting reflex is well documented (Hornby, 2001, Andrews and Horn, 2006, Horn, 2008), the precise brain mechanisms underlying nausea remain less clear. Nausea-induced conditioned gaping in rats has been shown to require an intact forebrain (Grill and Norgren, 1978b), and ablation of the insular cortex specifically prevents conditioned gaping (Kiefer and Orr, 1992), but not conditioned taste avoidance (Kiefer and Orr, 1992). These studies, therefore, suggest a critical role for the insular cortex in the sensation of nausea. Gustatory and visceral input (Cechetto and Saper, 1987, Allen et al., 1991) converge in the insular cortex with gustatory neurons occupying the anterior agranular and dysgranular layers (gustatory insular cortex [GIC]; Kosar et al., 1986), whereas visceral input converges on the posterior granular layer (visceral insular cortex [VIC]; Cechetto and Saper, 1987). Contreras et al. (2007) reported that LiCl-induced nausea resulted in VIC Fos-immunoreactivity, while inactivation of this region with lidocaine infusions reduced LiCl-induced lying on belly (LOB), a measure of unconditioned nausea in rats (Parker et al., 1984). Recently, work by our group (2012) demonstrated that the conventional anti-emetic, ondansetron, selectively blocked the establishment of conditioned gaping upon intra-VIC – but not GIC – administration, with the latter interfering with conditioned taste avoidance (Tuerke et al., 2012). Conversely, intra-VIC administration of the 5-HT3 agonist, mCPBG, potentiated LiCl-induced conditioned gaping and even produced conditioned gaping on its own (Tuerke et al., 2012). Limebeer et al. (2012) also showed a dissociation within the IC such that infusions of the cannabinoid agonist, HU210, directly into the VIC – but not GIC – interfered with the establishment of conditioned gaping, suggesting that the eCB system plays an important role within this forebrain region to modulate the sensation of nausea. The precise role of 2-AG and AEA in VIC regulation of nausea has remained unknown, however.
The current study investigated the function of the eCB system within the VIC, specifically, by assessing the roles of AEA and 2-AG-mediated signaling in this region and their potential to modulate nausea-induced conditioned gaping in rats. Separate experiments assessed whether exogenous 2-AG and AEA administration within the VIC were each capable of reducing conditioned gaping to investigate possible differences in eCB signaling during an episode of acute nausea, as well as determined whether the cannabinoid receptor type 1 (CB1) or downstream metabolites of 2-AG (e.g., Sticht et al., 2012) played a role in the suppressive effects of 2-AG.
Section snippets
Subjects
The subjects were male Sprague–Dawley rats weighing between 300 and 350 g at the start of experiments (Charles River Lab, St. Constant, Quebec). Animals were single-housed and maintained on a reverse light/dark cycle (7:00 am lights off; 7:00 pm lights on) with free access to food (Iams rodent chow, 18% protein) and tap water except during testing, which occurred during the dark cycle. All experiments were approved by the Animal Care Committee of the University of Guelph and were carried out in
Results
A photomicrograph of VIC cannulation (a) and schematic representation of the infusion cannula tip placements (b) for all rats is presented in Fig. 1. Cannulae were located between 0.00 and −1.32 mm posterior to Bregma. Dye spread analysis in a separate squad of rats (n = 14) revealed that micro-infusions of stain diffused within a range of 0.5–1.0 mm (mean = 0.65; SE = 0.04) A/P and 0.5–1.25 mm (mean = 0.82; SE = 0.06) M/L within the VIC; thus, despite the relatively large volume of infusate delivered in
Discussion
Here we report that pro-eCB manipulations within VIC selectively suppressed acute nausea-induced conditioned gaping in rats without interfering with avoidance of the nausea-paired taste during a two-bottle consumption test. Moreover, these anti-nausea effects appear to be mediated primarily by the eCB, 2-AG. In Experiment 1, animals were infused bilaterally with exogenous 2-AG, which was followed 15 min later by an injection of illness-inducing LiCl. It was found that the highest dose of 2-AG (1
Conclusions
The current study is the first published investigation of eCB modulation of nausea within the VIC, which is an important forebrain region involved in the sensation of nausea (Contreras et al., 2007, Limebeer et al., 2012, Tuerke et al., 2012). Although this study was limited to exogenous eCB administration, the results suggest that 2-AG may be the primary eCB modulating nausea within the VIC, and, overall, contributes to the emerging literature on the dissociable effects of 2-AG and AEA (see Di
Acknowledgments
The authors declare that there are no financial interests or conflicts of interest. The research was funded by operating grants from the Natural Sciences and Engineering Research Council of Canada (NSERC: 92057) and Canadian Institutes of Health Research (CIHR: 334086) to LAP, as well as an NSERC Canada Graduate Scholarship to MAS.
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