Retrograde Endocannabinoid Signaling Reduces GABAergic Synaptic Transmission to Gonadotropin-Releasing Hormone Neurons

Abstract

Cannabinoids suppress fertility via reducing hypothalamic GnRH output. γ-Aminobutyric acid (GABA)A receptor (GABAA-R)-mediated transmission is a major input to GnRH cells that can be excitatory. We hypothesized that cannabinoids act via inhibiting GABAergic input. We performed loose-patch electrophysiological studies of acute slices from adult male GnRH-green fluorescent protein transgenic mice. Bath application of type 1 cannabinoid receptor (CB1) agonist WIN55,212 decreased GnRH neuron firing rate. This action was detectable in presence of the glutamate receptor antagonist kynurenic acid but disappeared when bicuculline was also present, indicating GABAA-R involvement. In immunocytochemical experiments, CB1-immunoreactive axons formed contacts with GnRH neurons and a subset established symmetric synapses characteristic of GABAergic neurotransmission. Functional studies were continued with whole-cell patch-clamp electrophysiology in presence of tetrodotoxin. WIN55,212 decreased the frequency of GABAA-R-mediated miniature postsynaptic currents (mPSCs) (reflecting spontaneous vesicle fusion), which was prevented with the CB1 antagonist AM251, indicating collectively that activation of presynaptic CB1 inhibits GABA release. AM251 alone increased mPSC frequency, providing evidence that endocannabinoids tonically inhibit GABAA-R drive onto GnRH neurons. Increased mPSC frequency was absent when diacylglycerol lipase was blocked intracellularly with tetrahydrolipstatin, showing that tonic inhibition is caused by 2-arachidonoylglycerol production of GnRH neurons. CdCl2 in extracellular solution can maintain both action potentials and spontaneous vesicle fusion. Under these conditions, when endocannabinoid-mediated blockade of spontaneous vesicle fusion was blocked with AM251, GnRH neuron firing increased, revealing an endogenous endocannabinoid brake on GnRH neuron firing. Retrograde endocannabinoid signaling may represent an important mechanism under physiological and pathological conditions whereby GnRH neurons regulate their excitatory GABAergic inputs.

Marijuana, phytocannabinoids, the endocannabinoid system, and male fertility

“…As previously mentioned, cannabinoid receptors are closely related to neurons in the hypothalamus and GnRH release has been shown to be inhibited in males by AEA and THC through interaction with GABA and other systems [58,[80][81][82]. This reduction in gonadoliberins can cascade to the rest of the HPG axis as to be discussed subsequently (see Table 1).…”

“…This action could be prevented by a specific CB1 antagonist (SR141716) [56,57]. Farkas and coworkers furthermore demonstrated that endocannabinoid activates CB1 which, in turn, inhibits spontaneous gamma aminobutyric acid (GABA) release [58]. Postsynaptic GABA receptors, located on GnRH neurons, are not activated, and as a consequence, GnRH is not released.…”

“…Postsynaptic GABA receptors, located on GnRH neurons, are not activated, and as a consequence, GnRH is not released. Interestingly enough, the inhibitory effect of AEA was higher than that of 2-AG [58,59]. Olah and coworkers postulated that the difference between the effects of AEA and 2-AG on the serum levels of LH is due to the difference in receptor activation as AEA can activate both CB1 and TRPV1 receptors while 2-AG acts only on the CB1 receptor [60].…”

Distribution of type 1 cannabinoid receptor‐expressing neurons in the septal‐hypothalamic region of the mouse: Colocalization with GABAergic and glutamatergic markers

“…The presence of CB1 in this putative GABAergic input system to GnRH neurons may explain our recent electrophysiological and morphological data that retrograde endocannabinoid signaling reduces GABAergic afferent drive onto GnRH neurons via the activation of presynaptic CB1 receptors (Farkas et al, 2010). In that study we have provided evidence that GnRH neurons produce 2-arachidonoylglycerol (2-AG).…”

“…Its presynaptic action via CB1 decreases GABA release, which, in turn, reduces postsynaptic GABAA-R signaling and inhibits the firing activity of GnRH neurons (Farkas et al, 2010).…”