Abstract
Cholecystokinin (CCK) facilitates the process of satiation via activation of vagal afferent neurons innervating the upper gastrointestinal tract. Recent findings indicate CCK acts on these neurons via a ruthenium red (RuR) sensitive pathway that involves members of the vanilloid (V) subfamily of transient receptor potential (TRP) channels. To further test this mechanism, the mouse provides an ideal model in which genetic tools could be applied. However, whether CCK acts by similar mechanism(s) in mice has not been determined. In the present study we explored the actions of CCK on nodose neurons isolated from Sprague Dawley (SD) rat and two strains of mice; C57BL/6 and BalbC using fluorescence-based calcium imaging. With minor exceptions nodose neurons isolated from all species/strains behaved similarly. They all respond to brief depolarization with a large calcium transient. A significant subset of neurons responded to capsaicin (CAP), a TRPV1 agonist, although neurons from C57BL/6 were 10-fold more sensitive to CAP than SD rats or BalbC mice, and a significantly smaller fraction of neurons from BalbC mice responded to CAP. CCK-8 dose-dependently activated a subpopulation of neurons with similar dose dependency, percent responders, and overlap between CCK and CAP responsiveness. In all species/strains CCK-8 induced activation was significantly attenuated (but not completely blocked) by pretreatment with the TRPV channel blocker RuR. Surprisingly, the CCK analogue JMV-180, which is reported to have pure antagonistic properties in rat but mixed agonist/antagonist properties in mice, behaved as a pure antagonist to CCK in both rat and mouse neurons. The pure antagonistic action of JMV-180 in this in vitro preparation suggests that prior reported differential effects of JMV-180 on satiation in rats versus mouse must be mediated by a site other than vagal afferent activation.
Highlights
Coordination of behavioral and physiological responses following ingestion of food is critically dependent on neuronal transmission from the gastrointestinal (GI) tract to the brain [1]
This included a pure antagonistic action of JMV-180 in both rats and mice. These findings suggest that the mechanisms of CCK induced activation of vagal afferent neurons is conserved across species, and that mouse primary vagal afferent cultures may provide a useful tool in the dissection of CCK-1 receptor signal transduction pathways
Like the Sprague Dawley (SD) rat, cultured vagal afferents from BalbC and C57BL/6 mice were capable of maintaining low basal calcium levels for extended periods of time and exhibited rapid, robust, and reversible responses when challenged with high K+ (55 mM) containing baths, indicating viable expression of voltage-activated calcium channels and normal calcium sequestration pathways (Figure 1B)
Summary
Coordination of behavioral and physiological responses following ingestion of food is critically dependent on neuronal transmission from the gastrointestinal (GI) tract to the brain [1]. Release of the peptide cholecystokinin (CCK) from duodenal epithelium upon the arrival of nutrients into the duodenum activates vagal afferent terminals via CCK-1 receptors; a critical step in slowing gastric emptying, increasing pancreatic secretion, and facilitating the process of satiation [3]. GI projecting afferents provide key pre-absorptive nutritional information to the brain [4] and show enriched responsiveness to CCK [5]. In vagal afferents CCK acts via the low-affinity binding site [6,7,8,9] to decrease K+ [10,11] and increase non-selective cationic conductances [11], resulting in membrane depolarization and action-potential generation [9,12]. The specific cellular transduction pathway(s) and ionic conductances targeted by CCK binding at CCK-1 receptors remain incompletely characterized [13]
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