Roles of phospholipase Cbeta and NMDA receptor in activity-dependent endocannabinoid release

Abstract

Endocannabinoids are released from postsynaptic neurons, activate presynaptic cannabinoid receptors and cause various forms of short-term and long-term synaptic plasticity throughout the brain. Using hippocampal and cerebellar neurons, we have revealed that endocannabinoid release can be induced through two different pathways. One is independent of phospholipase Cbeta (PLCbeta) and driven by Ca(2+) elevation alone (Ca(2+)-driven endocannabinoid release, CaER), and the other is PLCbeta-dependent and driven by activation of G(q/11)-coupled receptors (receptor-driven endocannabinoid release, RER). CaER is induced by activation of either voltage-gated Ca(2+) channels or NMDA receptors. RER is functional even at resting Ca(2+) levels (basal RER), but markedly enhanced by a small Ca(2+) elevation (Ca(2+)-assisted RER). In Ca(2+)-assisted RER, PLCbeta serves as a coincidence detector of receptor activation and Ca(2+) elevation. We have also demonstrated that Ca(2+)-assisted RER is essential for the endocannabinoid release triggered by synaptic activity. Our anatomical data show that a set of receptors and enzymes required for RER are well organized so that the excitatory input can trigger RER effectively. Certain forms of spike-timing-dependent plasticity (STDP) are reported to depend on endocannabinoid signalling. The NMDA receptor and PLCbeta might play key roles in the endocannabinoid-dependent forms of STDP as coincidence detectors with different timing dependences.

Figure 1. Models of basal receptor-driven endocannabinoid release (RER) and Ca²⁺-assisted RER

A, at basal Ca²⁺ levels, strong activation of G_q/11-coupled receptors (e.g. mGluR1/5, M₁/M₃) stimulates PLCβ through Gα_q/11. PLCβ hydrolyses phosphatidylinositol 4,5-bisphosphate into diacylglycerol and inositol 1,4,5-trisphosphate. Diacylglycerol is then hydrolysed to 2-arachidonoylglycerol (2-AG) by diacylglycerol lipase (DAGL). 2-AG is released and activates presynaptic CB1 cannabinoid receptors, leading to the suppression of neurotransmitter release. B, when activation of G_q/11-coupled receptors coincides with Ca²⁺ increase through Ca²⁺-permeable channels (e.g. voltage-gated Ca²⁺ channel, NMDA receptor), PLCβ activation is enhanced. In this condition, 2-AG production can be induced even by weak activation of G_q/11-coupled receptors, which is subthreshold for basal RER.

Figure 2. Intracellular Ca²⁺ dependence of RER and PLCβ1-mediated TRPC6 activation driven by muscarinic activation

A, simultaneous whole-cell voltage clamp recordings were made from neuron pairs in rat hippocampal culture. The presynaptic neurons were stimulated and cannabinoid-sensitive IPSCs were recorded from the postsynaptic neurons dialysed with the solutions containing 30 mm BAPTA with the indicated pCa levels. Examples representing the effects of 2-AG (30 nm) and oxotremorine-M (oxo-M, 0.3 μm) at the four different pCa levels. IPSC traces acquired at the indicated time points are shown on the right. B, averaged data for oxo-M-induced suppression of IPSC at four different pCa levels buffered with 10–30 mm BAPTA or 10 mm EGTA. C and D, rat cultured hippocampal neurons expressing exogenous TRPC6 were dialysed with 10 mm BAPTA-containing solutions with the indicated pCa levels. Sample traces (C) and averaged amplitudes (D) of oxo-M (3 μm)-induced currents at the four different pCa levels. In the summary bar graphs (D), the amplitudes were normalized to the values for pCa 6. (Modified from Hashimotodani et al. 2005, with permission.)

Figure 3. Schematic drawings showing the molecular, anatomical and physiological organization for endocannabinoid signalling of the three cell types in the brain

A, in the Purkinje cell, diacylglycerol lipase-α (DAGLα) is densely concentrated at the base of spine neck. DAG produced at spine head diffuses to the base of spine neck and is converted to 2-arachidonoylglycerol (2-AG) by DAGLα. Released 2-AG activates CB1 cannabinoid receptors (CB1R) located on perisynaptic region of the parallel fibre (PF) terminal or nearby inhibitory terminal (In). B, in the hippocampal CA1 pyramidal cell, DAGLα is distributed in the spine head and neck. At this site, 2-AG is produced and travels to activate CB1R located on both excitatory (Ex) and cholecystokinin-positive inhibitory (CCK-In) terminals. The density of CB1R is low at the excitatory terminal, and high at the CCK-positive inhibitory terminal. PCD, Purkinje cell dendrite; PyD, pyramidal cell dendrite; PV-In, parvalbumin-positive inhibitory terminal. C, in the striatal medium spiny neuron, coincidental depolarization and mGluR5 activation are essential for PLCβ1/DAGLα-mediated production of 2-AG to induce retrograde suppression of corticostriatal synapse, because of low CB1 levels in excitatory corticostriatal afferents. This mGluR5-driven Ca²⁺-assisted RER is further facilitated with the aid of M₁, while M₁ activation alone fails to trigger 2-AG production because of its sparse distribution in the spines. This modulation will lead to the suppression of the hyperactivity of the medium spiny neuron (MSN). At MSN–MSN and parvalbumin (PV) interneuron–MSN synapses, which show high density of CB1R, both mGluR5 and M₁ can induce RER because of their widespread somatodendritic distributions. This retrograde suppression will lead to the increase of the excitability and striatal output of the MSN. (Modified from Yoshida et al. 2006 and Uchigashima et al. 2007, with permission.)