Opioid and endocannabinoid transmission is not just an “acute” signaling system related to pleasure, analgesia, or emotional regulation; it is, in fact, a modulatory system that, when activated repeatedly or chronically, is capable of shaping neuronal and glial gene expression through the activation of sophisticated and long-lasting intracellular signal transduction pathways. Functional impairments in these neurotransmission systems have been implicated in depression, depressive response and pain perception disturbances like chronic pain, fibromyalgia and others.
Effects of opioid transmission on gene expression
When β-endorphin or other opioid peptides bind the μ-opioid receptor (MOR), coupled to inhibitory G proteins (Gi/G0), a cascade of intracellular events occurs. In the acute phase, this leads to an inhibition of the membrane enzyme adenylate cyclase. It does not produce the second messenger cyclic AMP (cAMP) from ATP when inhibited by Gi or G0. This leads to the closing of calcium channels and opening of potassium channels with a hyperpolarizing and inhibitory effect on the release of various neurotransmitters. However, the most interesting effect for emotional plasticity and psychiatric disorders is the genomic and epigenetic one.
Chronic activation of opioid receptors triggers the compensatory activation of the protein kinase PKA and the transcription factor CREB; the latter facilitates the expression of genes associated with emotional resilience, such as the growth factor BDNF in the hippocampus and prefrontal cortex. In parallel, there is the silencing of genes related to the response to chronic stress (such as those encoding for glucocorticoid receptors). However, excessive, chronic or dysregulated exposure to opioid stimulation can determine negative epigenetic modifications, such as hyperacetylation of histones on DNA in limbic regions given by the excessive activation of protein kinases activated by extracellular stimuli (MAPK and calcium-dependent PKC).
This leads to increased expression of pro-depressive genes (e.g. those that increase the sensitivity of the amygdala to stress). When mu opioid receptors (MOR) in reward circuits are down-regulated, limbic hyporeactivity may appear with the phenomenon of secondary anhedonia, typical of major depression. In depressed patients, it has been observed, through post-mortem studies and molecular imaging, that the density and functionality of central opioid receptors is reduced in the prefrontal cortex and accumbens (Lutz et al., 2020).
Effects of endocannabinoid transmission on gene expression
Activation of CB1 and CB2 receptors by endocannabinoids such as anandamide (AEA) and 2-AG exerts an even more profound and pervasive genetic influence, which manifests itself through convergent and transverse signaling pathways. The CB1 receptor, abundantly expressed in the hippocampus, amygdala, prefrontal cortex and striatum, is coupled to Gi/G0 proteins and reduces cAMP production, thus reducing PKA function and phenomena regulated by it, such as synaptic vesicle release, energy production from glucose and cytoskeleton-induced cell shape changes.
But its prolonged activation reflexively modulates the ERK/MAPK (mitogenic protein kinase) pathway, because the beta-gamma subunits of the Gi or G0 protein can interact with the membrane protein kinase c-Src, which reaches the MAPKs via the c-Ras activator Sos1. CB1 activation stimulates, in the chronic phase, the Ras-Raf-MEK-ERK cascade, which promotes the transcription of neuroprotective and synaptic plasticity-associated genes, such as BDNF, EGF, CTNF, contactin-1, GPM6A, NCAM, S100B and Arc (Activity-Regulated Cytoskeleton-associated protein).
An important subsequent step is the coordinated expression of the nuclear factors Egr1-4, which serve to induce secondary target genes. Egr family encodes four zinc-finger transcription factors that are designated Egr1 (also known as zif268, NGFI-A, Krox24), Egr2 (also known as Krox20), Egr3 (also known as Pilot), and Egr4 (also known as NGFI-C). In neurons, Egr genes are robustly and transiently expressed by elevated cytosolic calcium produced by synaptic activity, and their expression is coupled with N-methyl d-aspartate (NMDA) receptor activation and MAPK signaling in excitatory glutamatergic synapses, by physiologic synaptic activity.
Arc (also known as Arg3.1) is a particularly interesting effector because, like Egr transcription factors, it is rapidly induced by synaptic activity, its expression depends upon NMDA receptor activation and intracellular MAPK signaling, and it has a critical role in maintaining LTP and long-term, but not short-term, memory formation. In activated neurons, Arc mRNA is rapidly distributed into dendrites, where it is targeted to recently activated synapses and locally translated into protein, suggesting that it has a direct role in the plasticity responses made within synapses after activation. Moreover, similar to Egr transcription factors, the Arc gene is regulated as an immediate early gene that is rapidly upregulated without the need for new protein synthesis.
Through the PI3K/Akt/mTOR pathway, endocannabinoid signaling regulates the local synthesis of synaptic proteins, facilitating processes of synaptogenesis and adaptation to the environmental context. This may be obtained by a parallel branching of the Src-Ras signaling: auto-phosphorylated c-Src couples directly withe p85 subunit of the PI3-kinase, which is also able to directly interact with membrane-bound K-Ras. PI3K then activates c-Akt which converge with ribosomal kinases (Rsk-1, Msk1) to regulare the mTOR complex toward the specific protein synthesis.
Interaction with epigenetic factors
CB1 activation modulates the activity of HDACs (histone deacetylases) and DNA methyltransferases, influencing chromatin structure and accessibility to genes that regulate anxiety and depression. This is directly obtained by the intervention of some protein kinases, namely ERK2, c-Akt, MSK-1. Human DNMT1 is discovered to be phosphorylated at Ser 154 by cyclin-dependent kinases (CDKs) 1, 2 and 5. Cdk5 is particularly important for neurons, since is not involved in cellular replication; rather, it controls cytoskeletal rearragements and, thus, synaptic remodeling.
AKT-mediated phosphorylation of DNMT1 at Ser143 peaks during DNA synthesis and stabilizes DNMT1 from degradation. Glycogen Synthase Kinase 3 (GSK3) phosphorylates DNMT1 at Ser714 to block the methylation of unmethylated DNA. The de novo DNA methyltransferase DNMT3a is also reported to be phosphorylated at two key residues (Ser386 and Ser389) by casein kinase 2 (CK2), which impairs the methylation ability of DNMT3a and switches DNMT3a to localize at heterochromatin (inactive DNA).
How this relates to anxiety and depression
Both opioid and endocannabinoid transmission play a critical role in regulating fronto-limbic circuits (amygdala, hippocampus, prefrontal cortex, nucleus accumbens), which are critical for managing anxiety and modulating mood. In anxiety disorders and major depression, it has been observed:
- Hypoproduction of brain endocannabinoids and reduced availability of CB1 (Hill et al., 2009)
- Reduced activity of the endogenous opioid system in cortical and limbic regions (Nummenmaa et al., 2020)
- Transcriptional dysregulation in genes associated with emotional resilience: decreased espression fo BDNF, Arc, Homer1a, FosB.
This creates a negative neurochemical cycle:
- Chronic stress → ↓ endocannabinoid and opioid tone
- ↓ Activation of ERK/MAPK, PKA-CREB pathways → ↓ favorable gene expression
- ↓ Neuroplasticity and ↑ vulnerability to anxiety and depression
Clinical and therapeutic implications
Molecular and genetic imaging studies suggest that interventions that restore the normal function of these systems (exercise, pharmacological, meditation, brain stimulation) induce favorable epigenetic changes, such as:
- ↑ Acetylation of histones in the hippocampus
- ↑ Transcription of BDNF and other synaptogenic genes
- ↓ Methylation of genes regulating plasticity
In practice, running, aerobic exercise, mindfulness and other natural stimuli are capable of “rekindling” these gene pathways and counteracting the molecular vulnerability to depression and anxiety.
- Edited by Dr. Gianfrancesco Cormaci, PhD, specialist in Clinical Biochemistry.
Scientific references
Nummenmaa L et al. Neuropsychopharmacol. 2020; 45(11):1953.
Morena M et al. Neuropsychopharmacol. 2016; 41(1):80.
Russo SJ, Nestler EJ. Nat Rev Neurosci 2013; 14(9):609.
McEwen BS, Morrison JH. Neuron 2013; 79(1):16-29.
Hill MN et al. Trends Pharmacol Sci. 2009; 30(9):484-93.
Li L et al. Mol Cell Biol, 2005; 25(23):10286–10300.
Guzowski JF et al. J Neurosci. 2000; 20:3993-4001.
