Excitability of prefrontal cortical pyramidal neurons is modulated by activation of intracellular type-2 cannabinoid receptors

Abstract

The endocannabinoid (eCB) system is widely expressed throughout the central nervous system (CNS) and the functionality of type-1 cannabinoid receptors in neurons is well documented. In contrast, there is little knowledge about type-2 cannabinoid receptors (CB 2 Rs) in the CNS. Here, we show that CB 2 Rs are located intracellularly in layer II/III pyramidal cells of the rodent medial prefrontal cortex (mPFC) and that their activation results in IP 3 Rdependent opening of Ca 2+ -activated Cl − channels. To investigate the functional role of CB 2 R activation, we induced neuronal firing and observed a CB 2 R-mediated reduction in firing frequency. The description of this unique CB 2 R-mediated signaling pathway, controlling neuronal excitability, broadens our knowledge of the influence of the eCB system on brain function.calcium-activated chloride current | firing rate | whole-cell current | voltage clamp | intracellular calcium stores T he endocannabinoid (eCB) system is involved in many functions of the CNS, including executive functions associated with the prefrontal cortex, such as decision-making and working memory (1). The eCB system consists of at least two G proteincoupled receptors (GPCRs), type-1 cannabinoid receptor (CB 1 R) and type-2 cannabinoid receptor (CB 2 R), lipid endogenous ligands (e.g., anandamide and 2-arachidonoylglycerol), and various enzymes responsible for the synthesis and degradation of the endogenous ligands (2-6). CB 1 Rs are among the most abundantly expressed GPCRs in the rat brain and their role, predominantly as presynaptic receptors, in modulating neurotransmission is clearly established (5,7,8). In contrast with CB 1 R, the presence and function of CB 2 R in the brain has long been a matter of debate (9). CB 2 Rs are found primarily in the immune system and were initially regarded as the "peripheral" cannabinoid receptor (10,11). This generally accepted idea is challenged by the description of CNS CB 2 R gene expression in rats and wild-type mice (12-14) and the identification of functional CB 2 Rs on glial cells and neurons (15)(16)(17)(18). In addition to the current view that supports the expression of functional CB 2 Rs in neurons upon brain stress or damage (19), it has been reported that CB 2 Rs could play a role in general CNS physiology (20)(21)(22). These developments emphasize the importance of understanding how CB 2 R activation affects neuronal functioning. To demonstrate the presence of functional CB 2 Rs in the rodent medial prefrontal cortex (mPFC) and to elucidate their functional role, we used Western blotting, a radioactive binding assay, and electrophysiological techniques (whole-cell current and voltage clamp) on layer II/III pyramidal neurons. ResultsFunctional CB 2 Rs in the mPFC. The presence of CB 2 Rs in the rat mPFC was demonstrated by a Western blot performed on homogenated mPFC samples (Fig. 1A). A band of the expected molecular weight for CB 2 R was detected, which was absent when the primary antibody was incubated with immunizing peptide (Fig. 1A)....

Cannabinoid CB 2 receptors modulate midbrain dopamine neuronal activity and dopamine-related behavior in mice

“…This inhibitory effect was reversed by pharmacologic blockade of CB 2 Rs (AM630) or was absent in CB 2 −/− mice, suggesting a CB 2 R-mediated effect. This finding is consistent with previous reports in which JWH133 or other CB 2 R agonists inhibited spontaneous and evoked neuronal firing to noxious stimuli in spinal cord and thalamus (44)(45)(46)(47) and inhibited excitatory neuronal firing in the prefrontal cortex (48). Thus, our electrophysiological data provide direct evidence demonstrating that the expressed CB 2 Rs in VTA DA neurons are functional, and that activation of these receptors inhibits VTA DA neuronal firing and decreases VTA DA neuronal excitability.…”

Differential Activation of Cultured Neonatal Cardiomyocytes by Plasmalemmal Versus Intracellular G Protein-coupled Receptor 55

“…Increases in intracellular Ca 2ϩ concentration as well as alterations in ionic currents and membrane properties have been correlated previously with GPR55 activation (16 -18). Considering the emerging paradigm of functional intracellular G protein-coupled receptors (GPCRs) (19,20), which has also been suggested for the receptors of other lipid mediators (21) and for cannabinoid receptors (22)(23)(24)(25), we postulated that a GPR55-dependent signaling pathway may be initiated either at the plasma membrane or * This work was supported, in whole or in part, by National Institutes of Health within the cell. Thus, we used Ca 2ϩ and voltage imaging and both extracellular and intracellular administration (microinjection) of GPR55 ligands to test the involvement of this receptor in the regulation of cardiomyocyte signaling.…”

G-protein Receptor Kinase 5 Regulates the Cannabinoid Receptor 2-induced Up-regulation of Serotonin 2A Receptors

“…CB 1 and CB 2 receptors bind endocannabinoids, synthetic cannabinoids, and cannabinoids found in nature (such as in Cannabis sativa) with high affinity (18,38). Although only CB 1 receptors were initially identified in the brain (39), later studies have also identified CB 2 receptors in several brain areas, including PFCx, hippocampus, amygdala, substantia nigra, and cerebellum (13,14), triggering a reevaluation of the possible roles that CB 2 receptors might play in the brain.…”

“…Accordingly, recent preclinical studies have indicated that chronic, but not acute, exposure to non-selective (10,11) or selective CB 2 receptor agonists induced anxiety-like behaviors in rodents (12). CB 2 receptors have been identified in postsynaptic neurons in several brain areas of the limbic brain, including brain areas such as the PFCx, hippocampus, and amygdala (13)(14)(15)(16). The CB 2 receptor is a prototypical G-protein-coupled receptor (GPCR) that couples to the G i/o class of G-proteins and can activate ERK1/2 signaling in either a G-protein-or ␤-arrestindependent pathway (17,18).…”