Excitatory neurotransmitters

Mechanism of action

Glutamate's action begins with its synthesis and storage in presynaptic neurons. The enzyme glutaminase catalyzes the conversion of glutamine from neighboring glial cells into glutamate. Additionally, glutamate can be synthesized through the transamination of 2-oxoglutarate, an intermediate of the tricarboxylic acid (TCA) cycle.

Once synthesized, glutamate is packaged into synaptic vesicles by vesicular glutamate transporters (VGLUTs). VGLUTs work as glutamate/proton antiporters: a vesicular H+-ATPase pumps protons into the vesicle lumen, and VGLUTs exploit this proton electrochemical gradient to drive glutamate uptake against its concentration gradient. Upon neuronal stimulation, a calcium (Ca²⁺) influx into the presynaptic terminal triggers glutamate release into the synaptic cleft through exocytosis.

Glutamate activity is rapidly terminated by reuptake into presynaptic neurons and glial cells via high-affinity excitatory amino acid transporters (EAATs). Within glial cells, the enzyme glutamine synthetase converts glutamate back into glutamine. This is essential for maintaining neuronal homeostasis and preventing excitotoxicity.

Glutamate receptors

There are two classes of glutamate receptors; ionotropic and metabotropic.

Ionotropic receptors (iGluRs) are ligand-gated ion channels that mediate rapid synaptic transmission. The three main subtypes:

AMPA receptors, which, upon glutamate binding, open as monovalent cation channels. Net Na⁺ influx and K⁺ efflux cause membrane depolarization. Standard AMPA receptors in the adult brain are not permeable to Ca²⁺, as the GluA2 subunit undergoes RNA editing that blocks calcium entry. Only GluA2-lacking AMPA receptors allow Ca²⁺ through, and these are restricted to specific plasticity contexts.

NMDA receptors, which are permeable to Na⁺ and K⁺, but especially Ca²⁺. Furthermore, due to the Mg²⁺ blockage at resting potential, NMDA receptors require both glutamate binding and membrane depolarization to be activated. Thus, they operate as coincidence detectors.

Kainate receptors, functioning similarly to AMPA. They remain less well understood.

Metabotropic receptors (mGluRs), are G-protein-coupled receptors (GPCRs) that act more slowly and modulate synaptic activity through intracellular signalling cascades, affecting gene expression and protein synthesis. Based on their pharmacological properties, mGluRs are divided into three classes:

Group I (mGluR1, 5): primarily postsynaptic and excitatory.

Group II (mGluR2, 3) and Group III (mGluR4,6,7,8): mainly presynaptic and inhibitory.

What does glutamate do?

Glutamate is the primary mediator of excitatory synaptic transmission. It plays a central role in neuroplasticity, which is essential for learning, cognition, and mood regulation. It contributes to processes like long-term potentiation (LTP). In regions such as the hippocampus and the cerebral cortex glutamate acting on NMDA and AMPA receptors strengthens synaptic connections and modulates dendritic spine density. Additionally, glutamate is important for motor control through its actions in the cerebellum and basal ganglia, where it contributes to movement initiation, planning, and coordination.

Mechanism of action

A highly efficient and tightly regulated process ensures effective communication between neurons releasing ACh and target cells. This includes the synthesis, storage, release, receptor binding and breakdown of ACh.

Choline, absorbed by cholinergic neurons via Na⁺-dependent uptake channels, combines with an acetyl group created from acetyl-CoA (produced in mitochondria) to form acetylcholine. This reaction is catalyzed by the enzyme choline acetyltransferase (ChAT).

In the axon terminal, acetylcholine is stored within synaptic vesicles via the vesicular acetylcholine transporter (VAChT). Once the action potential reaches the presynaptic terminal, it triggers the opening of voltage-gated Ca²⁺ channels, and, therefore, the influx of Ca²⁺ into the presynaptic neuron. This triggers the ACh-containing synaptic vesicles to fuse with the membrane via the SNARE complex, releasing ACh into the synaptic cleft through exocytosis, where it binds to its receptors.

To prevent overstimulation, after exerting its action, ACh is rapidly hydrolyzed in the synaptic cleft by an enzyme called acetylcholinesterase (AChE) into choline and acetate. Choline is then reabsorbed by the presynaptic neuron to participate in the next cycle of ACh synthesis.

Acetylcholine receptors

Once in the synaptic cleft, ACh can bind to either nicotinic or muscarinic receptors.

Nicotinic receptors (nAChRs) are ligand-gated, fast-acting ionotropic channels that promote rapid synaptic transmission by allowing Na⁺ and, to a lesser extent, Ca²⁺ to enter the postsynaptic cell, causing depolarization. They are further classified into two subtypes:

N1 (muscle-type), located at the NMJ on skeletal muscle cells playing a crucial role in muscle contraction.

N2 (neuronal-type), found in autonomic ganglia of the sympathetic and parasympathetic nervous systems, the adrenal medulla, and in various regions of the CNS including the ventral tegmental area, the prefrontal cortex, and the hippocampus.

Muscarinic receptors (mAChRs) are metabotropic GPCRs, initiating slower responses via intracellular signalling pathways. They are classified into five subtypes:

M1: excitatory, predominantly in the cerebral cortex, hippocampus, and gastric glands.

M2: inhibitory and found in cardiac tissue and smooth muscle, regulating heart rate and muscle relaxation.

M3: excitatory and involved in smooth muscle contraction and glandular secretion, as found in the smooth muscle cells of bronchioles, bladder, iris and small intestine.

M4 and M5: predominantly in the CNS and believed to play essential roles in processes related to cognition, reward, and mood regulation, though their function is not fully understood.

What does acetylcholine do?

ACh plays a vital role in both the CNS and PNS.

As the primary neurotransmitter at the NMJ, it ensures communication between motor neurons and skeletal muscle fibers. Once released, ACh binds to N1 nicotinic receptors on muscle cells, allowing Na⁺ to enter. This causes membrane depolarization, triggering muscle contraction- crucial for both voluntary and reflexive movements.

In addition to its role at the NMJ, ACh acts on muscarinic receptors to regulate a variety of autonomic processes. To maintain cardiovascular homeostasis, acetylcholine binds to M2 muscarinic receptors to mediate parasympathetic activity, reducing heart rate and cardiac contractility. In the gastrointestinal system, it promotes digestion by enhancing intestinal motility and stimulating secretion. ACh also induces bronchoconstriction via M3 receptors and triggers chemoreceptors, thereby influencing breathing reflexes. In the urinary system, it promotes bladder emptying by contracting the detrusor muscle, via M3 receptors mainly, and relaxing the internal urinary sphincter. Additionally, ACh supports thermoregulation and stimulates the secretion from exocrine glands, including the salivary, lacrimal, and sweat glands.

In the eye, it facilitates accommodation for near vision by contracting the ciliary muscle, and induces miosis (pupil constriction) by stimulating the sphincter pupillae muscle. Moreover, by parasympathetic pathways, it assists male sexual function, particularly by promoting penile erection through penile artery vasodilation. Another key role of ACh is the stimulation of the adrenal medulla via N2 receptors to release norepinephrine and epinephrine into the blood.

Finally, in the CNS, important cognitive functions like attention, learning, memory, and arousal are modulated by cholinergic projections.

Norepinephrine (NE)

Norepinephrine, also known as noradrenaline, is a fundamental catecholamine secreted by both neurons and adrenal glands. It is mainly produced by neurons in the locus coeruleus of the brainstem. These neurons project to all parts of the brain, including the cerebral cortex, thalamus, hypothalamus, olfactory bulb, midbrain, cerebellum, and spinal cord. Acting as both a neurotransmitter and a hormone, norepinephrine plays key roles in the fight-or-flight response, cardiovascular regulation, and attention.

Mechanism of action

Norepinephrine is synthesized from tyrosine, which is first converted into L-DOPA by the enzyme tyrosine hydroxylase. Subsequently, L-DOPA is decarboxylated via aromatic L-amino acid decarboxylase (AADC) to form dopamine, which is further converted into norepinephrine by the enzyme dopamine β-hydroxylase. This final step takes place within vesicles located at the adrenal medulla and sympathetic axon terminals, serving as the primary storage sites for norepinephrine. Upon stimulation, norepinephrine is released into the synaptic cleft or into the bloodstream where it can bind to adrenergic receptors. After exerting its effects, norepinephrine gets degraded either within the axon terminal or in the extracellular fluid by two key enzymes, monoamine oxidase (MAO) and catechol-O-methyltransferase (COMT).

Norepinephrine receptors

Norepinephrine acts on metabotropic adrenergic receptors, which are GPCRs. They are broadly classified into two main types: α (alpha) receptors and β (beta) receptors.

α-adrenergic receptors signal through different G-proteins; a1-receptors couple to Gq proteins and activate phospholipase C (PLC), which increases IP₃ and DAG, leading to elevated intracellular Ca², while a2-receptors are coupled to Gi proteins, which inhibit adenylyl cyclase, reducing cAMP levels.

α₁-receptors are primarily located on vascular smooth muscle and mediate vasoconstriction, thereby increasing blood pressure.

α₂-receptors are found presynaptically and function to inhibit the release of neurotransmitters, acting as part of a negative feedback mechanism.

β-adrenergic receptors are coupled to Gs proteins, which activate adenylyl cyclase, increasing cAMP and activating protein kinase A (PKA).

β₁-receptors are mainly located in the heart, where they increase heart rate and contractility.

β₂-receptors are found in smooth muscle, including in the lungs and blood vessels, where they mediate bronchodilation and vasodilation.

β₃-receptors are predominantly present in adipose tissue, where they promote lipolysis.

What does norepinephrine do?

In the CNS, norepinephrine plays a key role in modulating attention, memory and focus. It enhances the brain's reaction to stress and regulates the processing of sensory and cognitive inputs. Norepinephrine also prepares the body for immediate action by inducing vasoconstriction, which raises blood pressure and directs blood flow to vital organs and skeletal muscles. Furthermore, norepinephrine promotes oxygen intake by bronchodilation, increases heart rate and contractility, and affects key metabolic processes to assist with increased energy demand.

Epinephrine (EPI)

Epinephrine, also known as adrenaline, is a catecholamine hormone and neurotransmitter, primarily produced by the adrenal medulla and to a lesser extent by neurons. It is integral to the fight-or-flight response.

Mechanism of action

Epinephrine is synthesized from methylation of norepinephrine through the action of the enzyme phenylethanolamine N-methyltransferase (PNMT). This reaction occurs primarily in the cytosol of chromaffin cells in the adrenal medulla. The expression of PNMT is regulated by glucocorticoids, particularly cortisol, which is secreted by the adrenal cortex. Cortisol reaches the adrenal medulla via the bloodstream, where it upregulates PNMT expression, thereby enhancing epinephrine production. After its synthesis in the cytosol, epinephrine is transported back into secretory vesicles for storage until it is released into the bloodstream in response to stress or sympathetic stimulation. Upon release, epinephrine exerts its effects by binding to adrenergic receptors. It is eventually inactivated through enzymatic degradation, primarily by MAO and COMT, like norepinephrine.

Epinephrine receptors

Epinephrine like norepinephrine binds to α (alpha) and β (beta) adrenergic metabotropic GPCRs. It has a stronger affinity for β2-receptors, particularly in bronchial smooth muscle and skeletal muscle vasculature, where it promotes bronchodilation and vasodilation to optimize oxygen uptake and enhance blood flow. Epinephrine also activates β1-receptors in the heart to increase heart rate, contractility, and cardiac output. It also interacts with α1-receptors in vascular smooth muscle leading to vasoconstriction. Although it can bind to α2-receptors, its role in feedback inhibition of norepinephrine release is less prominent compared to norepinephrine.

What does epinephrine do?

By activating both α- and β-receptors, epinephrine functions as a powerful mediator of the fight-or-flight response. In the heart, it stimulates β1-adrenergic receptors, increasing the heart's contractility, and, therefore, enhancing cardiac output. Moreover, epinephrine induces pupil dilation (mydriasis) which can boost visual acuity during stress. Additionally, it promotes vasoconstriction via a1-receptors, raises blood pressure, and guarantees sufficient blood supply to vital organs such as the liver and skeletal muscles under stress. To meet the heightened energy demand, epinephrine stimulates glycogenolysis in the liver, providing an immediate source of glucose, and enhances lipolysis, increasing circulating free fatty acids for use as alternative fuel.

Mechanism of action

Dopamine's action depends on tightly regulated mechanisms involving synthesis, storage, release and receptor binding. Aromatic L-amino acid decarboxylase converts L-DOPA to dopamine in the cytoplasm of dopaminergic neurons. As already mentioned, dopamine may operate as a precursor for other catecholamines. Once synthesized, dopamine is transported into synaptic vesicles by the vesicular monoamine transporter 2 (VMAT2). Upon neuronal activation, exocytosis occurs, and dopamine is released into the synaptic cleft. Subsequently, it binds to dopamine receptors, D1-like or D2-like, on the postsynaptic membrane to exert its effects. Dopamine's activity is terminated by reuptake into the presynaptic neuron via the dopamine transporter (DAT) or enzymatic degradation by MAO and COMT.

Dopamine receptors

Dopamine receptors are GPCRs. There are five subtypes classified into two families based on structure and signaling pathways:

Excitatory D1-like receptors (D1, D5) increase cAMP by activating Gs or Golf proteins and enhance neuronal excitability, which is important for reward signaling and motor activity.

Inhibitory D2-like receptors (D2, D3, D4) reduce cAMP by coupling to Gi/o proteins and minimize neuronal responses. Presynaptic DA receptors (autoreceptors) primarily belong to the D2-like receptor family and regulate dopamine synthesis and release, serving as a feedback mechanism to prevent excessive DA activity.

What does dopamine do?

Dopamine drives reward signalling, motivation, and goal-directed behaviour. It also regulates motor control, working memory, and mood. For instance, anticipating or experiencing a pleasurable stimulus, such as a favorite treat, increases dopamine levels and reinforces the behavior. Beyond reward processing, dopamine is essential for learning, working memory, and attention. It also mediates control of movements, contributes to mood stability, and influences sleep and dreams.

Mechanism of action

The mechanism of serotonin action involves a series of tightly regulated steps, from its synthesis to its inactivation. Serotonin is synthesized from the amino acid tryptophan in a two-step enzymatic process; first, the tryptophan hydroxylase (TPH) converts tryptophan into 5-hydroxytryptophan (5-HTP). Then, aromatic L-amino acid decarboxylase (AADC) converts 5-HTP into serotonin. Once synthesized, serotonin is packaged and stored into vesicles by the VMAT2, thus protected from degradation and ready for release. Upon neuronal stimulation, serotonin is released into the synaptic cleft via Ca²⁺-dependent exocytosis. Subsequently, it either attaches to presynaptic autoreceptors, which regulate its release via negative feedback, or postsynaptic receptors, which mediate signal transmission. Serotonin is, then, removed from the synaptic cleft by the serotonin transporter (SERT) and recycled into presynaptic vesicles -a process called reuptake- or metabolized by MAO. Beyond the CNS, serotonin is also produced by enterochromaffin cells, metabolized in the liver, and occasionally transported to the lungs for further processing.

Serotonin receptors

Serotonin receptors, also known as 5-HT receptors, are a diverse group of receptors that mediate the effects of serotonin in the CNS and the PNS. They are grouped into seven major families, 5-HT1 through 5-HT7, which are further subdivided into at least 15 subtypes. Most 5-HT receptors are GPCRs, thus activating intracellular signaling cascades upon serotonin binding. One notable exception is the 5-HT3 receptor, a ligand-gated ion channel that mediates rapid neurotransmission.

5-HT receptors can be broadly classified as either excitatory or inhibitory:

5-HT1 receptors are primarily inhibitory. Activation of these receptors often reduces serotonin release through coupling with Gi/o proteins, which inhibit adenylyl cyclase and decrease cAMP levels. They are often found presynaptically as autoreceptors.

5-HT2 receptors generally exert excitatory effects. They are coupled to Gq proteins, which increase synaptic activity by inducing intracellular signaling pathways such as PLC activation, leading to elevated intracellular Ca²⁺ and enhanced neuronal excitability.

5-HT3 receptors form ligand-gated ion channels that mediate fast excitatory neurotransmission.

5-HT4, 5-HT6, and 5-HT7 receptors are primarily excitatory and are all coupled to Gs proteins. Activation of these receptors stimulates adenylyl cyclase, leading to increased cAMP production.

What does serotonin do?

Serotonin's actions influence vital processes like mood regulation, learning, appetite, hemostasis, sleep, and memory.

In the CNS, serotonin contributes to mood stabilization and mental health regulation. It is also significant for neuroplasticity, assisting in learning and memory formation, and for cognitive functions like attention and decision-making. Moreover, serotonin affects the hypothalamus, aiding in thermoregulation and hunger regulation. In the brainstem, it modulates cardiovascular and respiratory rhythms.

In the periphery, approximately 90% of the body's serotonin is stored in enterochromaffin cells of the gastrointestinal tract. There, it regulates intestinal motility. At high concentrations, serotonin induces vasoconstriction, while at lower concentrations, it promotes vasodilation.

Mechanism of action

Histamine is synthesised intracellularly from the amino acid L-histidine by histidine decarboxylase (HDC), a pyridoxal-5'-phosphate-dependent enzyme that removes the carboxyl group from histidine to produce histamine. The main factor controlling the synthesis of histamine is the availability of histidine. Once synthesized, histamine is stored in vesicles until released in response to various stimuli. Its release is strictly modulated by H3 autoreceptors through negative feedback as well histamine N-methyltransferase (HNMT), and MAO-B through enzymatic degradation. This precise regulation ensures balanced control over wakefulness, neuronal excitability and circadian rhythms. Beyond the CNS, histamine is also released from enterochromaffin-like cells in the stomach and from immune cells, where it plays key roles in gastric acid secretion and immune responses, respectively.

Histamine receptors

Histamine receptors are metabotropic GPCRs and they comprise four subtypes:

H1 receptors are widely distributed throughout the body and brain (smooth muscle, endothelium, CNS). They are coupled to Gq proteins and activate signaling pathways leading to Ca²⁺ release. They mediate allergic responses, vasodilation, bronchoconstriction, itching, and wakefulness.

H2 receptors are primarily excitatory, and expressed in gastric parietal cells, the heart and the CNS. They are coupled to Gs proteins and increase intracellular cAMP. Their activation leads to gastric acid secretion, cardiac stimulation, and smooth muscle relaxation.

H3 receptors function as autoreceptors in the brain, regulating histamine production and release, and as heteroreceptors modulating other neurotransmitters. They are coupled to Gi/o proteins, inhibit adenylyl cyclase, reduce cAMP, and modulates Ca²⁺/K⁺ channels.

H4 receptors are found mainly in immune cells (eosinophils, mast cells). They largely regulate immune cell chemotaxis, inflammatory responses, and may contribute to allergic conditions. They are coupled to Gi proteins.

Histamine influences ionotropic receptors indirectly, likely through interactions with histamine-sensitive GABA receptor subunits, and also enhances NMDA receptor activity, a mechanism connected to synaptic plasticity and neuronal signaling.

What does histamine do?

In both the CNS and PNS, histamine is essential for the regulation of numerous physiological functions. In the brain, it is important for maintaining wakefulness, attention, and circadian rhythms, as well as memory modulation, sleep-wake transitions, and cognitive functions. Particularly in areas such as the hippocampus and amygdala, it affects motor coordination, emotional reactions such as fear, synaptic plasticity, and long-term memory consolidation. On the periphery, histamine is involved in immune responses, inflammation, allergic reactions, and it helps regulate pain perception and gastric acid secretion, thereby supporting gastrointestinal homeostasis.