Hallucinogens and Serotonin 5-HT2A Receptor-Mediated Signaling Pathways

This book chapter (2018) describes the history, physiological response to psychedelics, and signaling pathway activated by serotonin (5-HT) 2A receptors.

Abstract

“The neuropsychological effects of naturally occurring psychoactive chemicals have been recognized for millennia. Hallucinogens, which include naturally occurring chemicals such as mescaline and psilocybin, as well as synthetic compounds, such as lysergic acid diethylamide (LSD), induce profound alterations of human consciousness, emotion, and cognition. The discovery of the hallucinogenic effects of LSD and the observations that LSD and the endogenous ligand serotonin share chemical and pharmacological profiles led to the suggestion that biogenic amines like serotonin were involved in the psychosis of mental disorders such as schizophrenia. Although they bind other G protein-coupled receptor (GPCR) subtypes, studies indicate that several effects of hallucinogens involve agonist activity at the serotonin 5-HT2A receptor. In this chapter, we review recent advances in understanding hallucinogen drug action through characterization of structure, neuroanatomical location, and function of the 5-HT2A receptor.”

Authors: Juan F. López-Giménez & Javier González-Maeso

Notes

This paper is included in our ‘Top 12 Articles on Psychedelics and Serotonin (5HT) Receptors‘

Summary

The neuropsychological effects of naturally occurring psychoactive chemicals, such as mescaline and psilocybin, have been recognized for millennia. Recent studies indicate that several effects of hallucinogens involve agonist activity at the serotonin 5-HT2A receptor.

1 Introduction to Receptor Pharmacology

The concept of cellular receptors was originally conceived at the end of the nineteenth century and the beginning of the twentieth century by pioneering physiologists such as Langley and Erlich. Alfred Joseph Clark introduced the receptor theory in the 1930s.

Several forms of receptors exist, including enzymes, ion channels, and transporters, as well as nuclear and cytosolic proteins. Seven transmembrane (7TM) receptors are the largest group of these receptors, and they are peptides that cross the lipid bilayer through seven transmembrane domains.

The most extensively characterized mechanism of action of these receptors resides in their coupling to G proteins, which in turn initiate different second messenger cascades at the intracellular level.

All cell signaling processes, initiated upon the activation of GPCRs, can be terminated at different points. These include the hydrolysis of bound GTP to GDP by the G subunit, and the phosphorylation of different amino acids in the intracellular loops or carboxy terminus by specific receptor kinases.

Recent investigations have revealed that -arrestins play new roles in addition to those originally linked to receptor endocytosis, and that -2 adrenoceptors can couple to G subunits from the lumen of endosomes, meaning that the activation of intracellular effectors continues following endocytosis. Approximately 900 different types of GPCRs have been identified in the human genome, and are the therapeutic target of nearly half of all medicines.

2.1 Serotonin

Serotonin was discovered in the middle of the nineteenth century and was named after the compound that caused smooth muscle contraction. It was later synthesized and found to have the same properties as the substances obtained from natural sources.

During the period of intense research activity that coincided with the discovery of 5-HT, it was hypothesized that the hallucinogenic properties of LSD were mediated by its interaction with the serotonergic system in the CNS.

5-HT is present in all animal organisms and participates in numerous physiological functions. It is located preferentially in the brainstem.

5-HT is synthesized from tryptophan, an essential amino acid, following two biochemical reactions. The principal route of degradation of 5-HT is by deamination, which is performed by monoamine oxidase (MAO) enzymes.

5-HT is a neurotransmitter that is synthesized in the brainstem and releases from terminals to interact with specific receptors located mainly in postsynaptic neurons. 5-HT is either stored for future synaptic release or metabolized by MAO enzymes.

5-HT binds to 14 different cell membrane receptors, including 5-HT3 receptors, which belong to the ion channel receptor superfamily.

2.2 Serotonin Receptors

Several different 5-HT receptor subtypes were pharmacologically characterized in mammalian brain homogenates using radioligand binding techniques newly developed during the mid-1970s. These studies differentiated between two classes of 5-HT sites: 5-HT1 receptors and 5-HT2 receptors. The 14 5-HT receptor subtypes that are currently known were cloned using new molecular biology techniques. They are classified based on their primary structure, which is determined by their amino acid sequence.

The 5-HT2 receptor subfamily comprises three different subtypes, namely 5-HT2A, 5-HT2B and 5-HT2C, and they are grouped together due to their high structural homology. The 5-HT2A receptor is responsible for the neuropsychological effects of serotonergic hallucinogens.

3 Chemical Neuroanatomy of 5-HT2A Receptors

Hallucinogens induce altered states of consciousness, which are mostly generated by the interaction of hallucinogens with 5-HT2A receptors.

Hallucinogens bind to 5-HT2A receptors and activate them to generate hallucinations and a behavioral response. The anatomic distribution of 5-HT2A receptors in the CNS is essential to understand these effects. Radiochemicals based on the isotopic labeling of particular ligands can be used to visualize the distribution of receptors in the human brain. Initial studies using radioligands to study 5-HT2A receptor localization were not completely selective because they bind to sites corresponding to dopamine receptors, adrenergic receptors, or other 5-HT2 receptor subtypes. The development of [3H]MDL100,907, a highly selective 5-HT2A receptor antagonist, addressed many of the problems associated with other 5-HT2A receptor radioligands.

The anatomic localization of 5-HT2A receptors in primate brain showed a heterogeneous and wide-ranging distribution throughout different brain areas, with the highest density being in the neocortex. Initial investigations of 5-HT2A receptor distribution in the brain showed a homogeneous pattern with a high component of nonspecific binding, but when agonist radioligands were used, the pattern was markedly heterogeneous.

Striosomes are anatomical structures differentiated from the rest of the striatum or matrix. They receive inputs from limbic regions and project to the pallidum and substantia nigra pars reticulata, whereas matrix afferents target dopaminergic nigral neurons.

Human, mouse, guinea pig, and cow brain all contain 5-HT2A receptors in striosomes, whereas no striosome labeling has been detected in rat, cat, pig, cow, or monkey brain. This difference may have fundamental functional consequences when evaluating drugs that interact with 5-HT2A receptors.

4. Hallucinogenic and Non-Hallucinogenic 5-HT2A Receptor Agonists

The mechanism of action of hallucinogens has been studied by pharmacologists and neuroscientists for decades. The role of the 5-HT2A receptor in the mechanism of action of hallucinogens was first proposed by Richard Glennon, Milt Titeler and their teams in 1984, 1986.

From a basic pharmacological perspective, it is particularly interesting that certain closely related 5-HT2A receptor agonists do not behave as hallucinogens in humans, but are used as therapeutic drugs in the treatment of migraine and Parkinson’s disease.

Individual GPCRs can couple to multiple signal transduction pathways, and agonists can stabilize distinct active conformational receptor states. These active states can elicit different patterns of cellular signaling responses.

The ternary complex model, which was first proposed by Robert Lefkowitz and his group, proposes that GPCRs are in a dynamic equilibrium between the inactive (R) and the active (R*) conformational states, and that agonists have higher affinity for the active state.

Terry Kenakin proposed a model where GPCRs adopt multiple conformational states, and different agonists show a preference for a subset of these states. This model could be used to design new drugs that specifically affect specific signaling pathways.

Kelly Berg, William Clarke, and their team found that different serotonin 5-HT2A receptor agonists activate different signaling pathways, which may explain the different neuropsychological effects of hallucinogenic and non-hallucinogenic 5-HT2A receptor agonists.

5 Biased Agonism at the 5-HT2A Receptor

The 5-HT2A receptor-coupled signaling pathway is activated by Gq/11, leading to inositol phosphates and diacylglycerol, followed by Ca2+ release from the endoplasmic reticulum. However, whether this pathway plays a role in mediating the effects of hallucinogens is uncertain.

Studies in heterologous expression systems and in mouse models suggest that several signaling cascades are involved in hallucinogen-induced behavioral response, including Gq-dependent signaling, PLA2-mediated arachidonic acid release, and Gi/o-dependent G-associated activation of ERK1/2.

Signaling pathways regulate gene expression in response to extracellular stimuli, and hallucinogenic and non-hallucinogenic 5-HT2A receptor agonists modulate specific signaling pathways that are responsible for their behavioral effects. Both hallucinogenic and non-hallucinogenic drugs modulate neuronal signaling in somatosensory cortex via the 5-HT2A receptor. Additionally, the entire transcriptome fingerprint induced by the hallucinogenic and non-hallucinogenic agonists was abolished in 5-HT2A knockout mice.

Hallucinogens modulate both Gq/11 and Gi/o proteins in mouse cortical primary cultures, and this explains how hallucinogens produce different neurophysiological responses to the same population of cortical pyramidal 5-HT2A receptors.

In HEK293 cells, hallucinogenic and nonhallucinogenic 5-HT2A receptor agonists induce distinct patterns of protein phosphorylation. Hallucinogens selectively activate Gi/o-dependent signaling, whereas non-hallucinogenic 5-HT2A receptor agonists do not activate Gi/o.

The role of -arrestins in GPCR function is further supported by the recent crystal structure of active -arrestin-1 bound to a -derived carboxyl terminal, and by the fact that 5-HTP and DOI activate a signaling cascade composed of -arrestin-2, phosphoinositide 3-kinease, Src, and Akt.

Recent findings have shown that the 5-HT2A receptor contains a PDZ-binding domain. This domain is potentially involved in the unique behavioral effects induced by hallucinogens.

The 5-HT2A receptor is associated with protein networks that are important for its synaptic localization and coupling to signaling machinery. The association of the 5-HT2A receptor with PSD-95 enhances 5-HT2A receptor-dependent Gq/11-coupling and inhibition of agonist-induced 5-HT2A receptor internalization.

The cytokines and growth factors that induce cell growth and contraction are mediated by the evolutionary conserved Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathway. The hallucinogen DOI activates both the MEK-ERK1/2 and JAK2-STAT3 intracellular signaling pathways.

The N376PxxY motif found at the junction between TM7 and the carboxyl terminal domains of the 5-HT2A receptor is essential for ADP-ribosylation factor (ARF)-mediated signaling, and ARF1 rather than ARF6 participates in this mechanism through a GTP-dependent interaction with the carboxyl terminus of the 5-HT2A receptor.

GFP-tagged 5-HT2A receptors were shown to internalize in the absence of 5-HT and to recycle to the cell surface in the presence of 5-HT. The human 5-HT2A receptor takes longer to recycle to the cell surface than the rat 5-HT2A receptor.

Genetic deletion of p90 ribosomal S6 kinase 2 (RSK2) potentiates 5-HT2A receptor-dependent signaling, as demonstrated in studies of fibroblasts obtained from wild-type and RSK2 knockout mice.

Several lines of evidence have shown that 5-HT1A and 5-HT2A receptors often show opposite effects on common signaling pathways, and that 5-HT2A receptors attenuate the effect of 5-HT1A receptors on NMDA receptor currents and microtubule depolymerization.

DARPP-32 is a key regulator of kinase-phosphatase signaling cascades modulated by serotonergic, dopaminergic, and glutamatergic neurotransmission. Three pathways regulate the state of phosphorylation of DARPP-32, which leads to increased phosphorylation of various PP1 substrates, which are involved in the psychoactive behavioral effects of these drugs.

DOI induces a differential regulation of BDNF mRNA expression in rat hippocampus and neocortex, which is blocked by selective 5-HT2A antagonists but is unaffected by selective 5-HT2C antagonists. Additionally, mGlu2/3 receptor agonists may modulate 5-HT2A receptor-dependent stress-induced behaviors.

7 Role of mGlu2 Receptor in Hallucinogen Action

Although 5-HT2A receptors are the main molecular target responsible for the cellular, electrophysiological and behavioral effects of hallucinogens in rodents, mGlu2/3 receptors may also contribute. This is demonstrated by the antagonization of 5-HT2A receptor-induced excitatory postsynaptic potential/currents in pyramidal neurons.

GPCRs are thought to function as monomers, but many instances of homomerization and heteromerization have been reported. This is further suggested by the recent explosion of research elucidating the crystal structures of GPCRs. Previous findings suggest that the Gq/11-coupled 5-HT2A receptor and the Gi/o-coupled mGlu2 receptor form a specific GPCR heteromeric complex in mouse and human frontal cortex. This complex is supported by the co-expression and co-immunoprecipitation of these receptors in cortical synaptic junctions.

The 5-HT2A-mGlu2 receptor complex is critical for the hallucinogen-like behaviors induced by 5-HT2A receptor agonists, and the expression of 5-HT2A and mGlu2 receptors in the frontal cortex of schizophrenic subjects postmortem supports this conclusion.

8 Future Directions

Recent work suggests that hallucinogens modulate the activity of specific 5-HT2A receptor-linked signaling pathways, and may be used as therapeutic drugs for the treatment of severe psychiatric and neurological disorders.

Fig. 1.

Chemical structures of hallucinogen and non-hallucinogen 5-HT2A receptor agonists, and competition curves for ketanserin by LSD, mescaline, lisuride and ergotamine in mouse somatosensory cortex plasma membrane preparations.

Fig. 3.

The ternary complex model describes how the receptor undergoes a conformational transition to an active state and interacts with heterotrimeric G proteins.

Fig. 5.

Immunogold labeling for 5-HT2A and mGlu2 receptors in mouse cortical neurons shows that the receptors are located in very close proximity at the synaptic junction. mGlu2 expression is necessary for head-twitch psychosis-like behavior induced by hallucinogenic 5-HT2A agonists.