28.9: Gated Ion Channels - Neural Signaling

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Learning Goals 

 

(Learning goals written by Claude, Sonnet 4.6, Anthropic)

Membrane Potentials, Ion Gradients, and the Action Potential

• Explain how the Na⁺-K⁺-ATPase establishes transmembrane ion gradients and contributes to the resting membrane potential (~-65 mV in neurons), and describe how differential membrane permeability to K⁺, Na⁺, and Cl⁻ through non-gated and gated channels converts these chemical gradients into an electrical potential.
• Trace the sequence of ionic events during an action potential: ligand-gated receptor depolarization → voltage-gated Na⁺ channel opening and Na⁺ influx → peak depolarization → voltage-gated K⁺ channel opening and K⁺ efflux → hyperpolarization → return to resting potential, and explain the structural basis for selectivity in the Na⁺ and K⁺ channels, including the role of the selectivity filter and dehydration of ions.
• Distinguish between ionotropic receptors (ligand-gated ion channels producing fast, millisecond-scale responses) and metabotropic receptors (GPCRs that indirectly modulate ion channels through second messengers, producing slow, seconds-to-minutes responses), and give examples of neurotransmitters that act through each.

Inhibitory Neurotransmission and the GABA Receptor

• Describe the subunit composition and architecture of the GABA_A receptor, explain how GABA binding opens a Cl⁻-selective channel to hyperpolarize the postsynaptic membrane, and explain the pharmacological mechanisms by which benzodiazepines (positive allosteric modulators), picrotoxin (channel blocker), and ethanol (indirect agonist) each modify receptor function at distinct binding sites.
• Explain why the combination of ethanol and benzodiazepines is synergistically lethal, and describe the mechanism by which Ro-15-4513 acts as a partial inverse agonist to antagonize both ethanol and benzodiazepine effects on the GABA_A receptor.

Metabotropic Signaling, Psychoactive Drugs, and Neuroplasticity

• Explain how metabotropic GPCRs in the brain (serotonin, dopamine, adrenergic, adenosine receptors) modulate neuronal excitability through second messenger cascades that phosphorylate ion channels, regulate gene expression via CREB, or promote receptor desensitization—using caffeine's blockade of the adenosine A2A receptor as a concrete example.
• Describe how LSD, psilocybin/psilocin, and serotonin differ in their binding to the 5-HT2A receptor (orthosteric site vs. extended binding pocket), explain the hypothesis that lipophilicity-dependent access to intracellular GPCRs on organelle membranes may contribute to hallucinogenic effects, and contrast this with non-hallucinogenic neuroplasticity-promoting mechanisms.
• Explain how both classical antidepressants (SSRIs, tricyclics) and psychedelics (LSD, psilocin) promote neuroplasticity by binding to the transmembrane domain of the TrkB/BDNF receptor, and describe the structural basis for why LSD binds with ~1000-fold higher affinity than fluoxetine through distinct but overlapping binding modes within the TrkB TMD dimer interface.

Introduction to Neural Signaling

Ion channels are membrane proteins that allow the flow of normally impermeant ions across the hydrophobic "sea" of cell or organelle membranes. The channel proteins can be constitutively open or "gated" open (or closed) by signals that affect the protein's conformation. The signals can be ligands (such as hormones or neurotransmitters), post-translational modifications (mostly phosphorylation and dephosphorylation), or more complex events (changes in transmembrane potential, pressure, or temperature). Gated channels are perhaps best studied in terms of their role in neural signaling.

Neurochemistry is one of the most explosive areas of biological research. Scientists are unraveling the molecular bases for memory, cognition, emotion, and behavior. The next decades will bring a greater understanding of brain chemistry, along with it the potential to alter human mood and memory, and to treat mental illnesses such as schizophrenia much more effectively. The human brain has about 86 billion neurons. Compare this with estimates for the number of stars in the Milky Way galaxy (about 200 billion, derived from luminosity and mass measurements, but some estimates suggest 400 billion). Now imagine that each neuron can form connections - synapses - with 1000 to 10,000 other neurons. Throw in another 86 billion nonneuronal cells, like glial cells, and you have one of the most complex structures in the universe. A recent high-resolution map of the cerebral cortex shows that a single cubic millimeter of the brain contains 57,000 cells, about 150 million synapses (connections between cells), and about 0.2 meters of blood vessels.  This section will explore the biology and chemistry of neurons.

Recent studies suggest that there are around 3,300 different types of cells in the brain.  We will discuss two kinds of neurons - those that interact with muscles at the neuromuscular junction and those that interact with other neurons in the central nervous system. Neurons consist of a single, nucleated cell body with multiple signal-receiving outgrowths (short, branched dendrites for receiving electrical impulses) and multiple signal-sending outgrowths (long, single axons for sending electrical impulses, myelinated for insulation) that end in terminal buttons.

These interact at synapses with the dendrites of other neurons. These characteristics are shown in Figure \(\PageIndex{1}\).

Figure \(\PageIndex{1}\): A neuron. Bruce Blaus - Own work, CC BY 3.0, https://commons.wikimedia.org/w/inde...curid=28761830

The high-resolution map of the human cortex shows that an axon most commonly (96%) forms one synapse with a target cell.  3% of the axons form 2 synapses with the target cell.  However, neurons that had lots of input onto their dendrites (they were highly innervated with more than 3000 synapses to their dendrites) could form 7 or more synapses from their axon to a target cell. 

A presynaptic neuron can stimulate an adjacent postsynaptic neuron by releasing a neurotransmitter into the synapse between the cells, which binds to a receptor in the membrane of the postsynaptic cell, stimulating the cell, as shown in Figure \(\PageIndex{2}\).

Figure \(\PageIndex{2}\): The synapse. https://cnx.org/contents/FPtK1zmh@8.25:fEI3C8Ot@10/Preface..Creative Commons Attribution 4.0 International license.

We will discuss the events that cause the post-synaptic cell to "fire," but we will not discuss the immediate events that lead to the presynaptic neuron's release of neurotransmitters.

Neurons (like all cells) have a transmembrane potential across the membrane. Transient changes in the membrane potential are associated with neuron activation or inhibition. This arises partly from the ionic imbalance across the membrane, which the Na⁺-K⁺-ATPase establishes. This electrogenic antiporter transfers 3 Na ions out of the cytoplasm for every 2 K ions it transports in. Likewise, Cl^- has a much higher level outside the cell. Membrane potentials are determined by the size of the ion gradients across the membrane and by the differential permeability of membranes to ions. As we saw previously, synthetic bilayer membranes are not very permeable to ions, as shown in Table \(\PageIndex{1}\) below.

Ion permeability of phosphatidyl serine vesicles

ION	PERMEABILITY (cm/s)
sodium	<1.6 x 10^-13 (lowest)
potassium	<9 x 10^-13
chloride	<1.5 x 10^-11 (highest)

Table \(\PageIndex{1}\): Ion permeability of phosphatidyl serine vesicles

Sodium would be expected to have a lower permeability than potassium since it has a higher charge density. It is also the largest in effective size due to the larger hydration sphere around the ion arising from its higher charge density. Chloride would be expected to have the highest permeability because it has the lowest charge density (due to electron cloud repulsion in the negatively charged ion). Intracellular charged proteins (which are mostly negative) are not permeable and help create the negative charge imbalance across the membrane.

Much work has been done on the giant axon of the squid, which has uniquely high intracellular potassium (400 mM vs 20 mM outside), high extracellular sodium (440 mM vs 50 mM inside), and chloride (560 mM vs 52 mM inside). Mammalian cell concentrations are much lower, but the relative size of the gradient is about the same. Typical ion concentrations and permeabilities for mammalian membranes are shown in Table \(\PageIndex{2}\) below.

T

Ion	Cell (mM)	Blood (mM)	Permeability (cm/s)
potassium	140	5	5 x 10^-7
sodium	5-15	145	5 x 10^-9
chloride	4	110	1 x 10^-8
X^- (negative macromolecules)	138	9	0

Table \(\PageIndex{2}\): Typical ion concentrations and permeabilities for mammalian membranes

How can we account for the markedly greater permeabilities of ions (1000x to 1,000,000x) in mammalian cell membranes compared to synthetic lipid vesicles? Previously, we showed that glucose has greater permeability through red blood cell membranes than through synthetic liposomes because of a membrane receptor that enables facilitated diffusion down its concentration gradient. The same holds true for ion permeabilities in intact biological membranes. These membranes have different selective ion channels (non-gated - always open, and gated - open only after specific conformational changes). The non-gated channels dramatically increase membrane permeability to ions, whereas the glucose transport protein increases permeability to glucose. It turns out that this differential permeability contributes to the transmembrane potential. Ion channels in nerves and muscles can move ions across the membrane at a rate of up to 10⁹/s, comparable to k_cat for the best enzymes.

If we envision channels as pores, how can we account for their selectivity for specific ions? A larger pore should admit any ion less than a maximal size for the pore, so it is hard to imagine the nature of the selectivity filter. Because of this, many people discounted the idea of channels in favor of a transporter, which would bind the ion selectively and then, through conformational changes, move the ion across (much like the Na⁺/K⁺ ATPase we discussed in the previous guide). This model, however, could not account for the incredible rates of ion flux across the membrane. Selectivity can be accounted for by a channel that contains a narrow opening that acts as an ion sieve. The ion loses most of its hydration sphere and forms specific interactions with amino acid side chains in the pore region. Such an interaction would be transient and not too tight since the ion must pass through the membrane. As we will see later, these ion channels:

• pass ions down a concentration gradient in a thermodynamically favorable process
• are specific for certain ions (although a few are less selective and will pass Na, K, Ca, and Mg ions)
• allow ion flow through either ungated or gated channels
• saturate with increasing ion concentration (even though concentration increases, the ions have a greater thermodynamic drive to pass through the channel). This is consistent with the ion binding at a selectivity filter in the narrow part of the pore. The K_D for the interaction is usually in the mM range and indicates weak binding with large dissociation rate constants (k_off).

Transmembrane Potentials

Several questions arise regarding the distribution of ions and the magnitude of the transmembrane potential.

1. How are the ion gradients established?
2. How does the transmembrane ion distribution contribute to the membrane potential?
3. How can the resting electrochemical potential and the ion distribution be maintained?

The answer to these questions will be illustrated using studies on two types of brain cells, glial cells (which function as protectors, scavengers, and feeders for brain neurons) and neurons. Both types of cells have transmembrane potentials.

Glial Cells

Different mechanisms can establish transmembrane ion gradients. One uses ion-specific ATPases (P-type ion transporters), as we discussed with the Na/K ATPase. This transporter ejects three sodium ions from inside the cell for every two potassium ions it transports in, all against their respective concentration gradients. As an electrogenic antiporter, it helps generate the potential. Specific ion channels also contribute (as described below) to the transmembrane gradients and potentials.

The harder question is how the distribution of ions contributes to the membrane potential. Two things must occur for a membrane potential to exist: First, there must be a concentration gradient of charged ions (for example, sodium, potassium, or chloride) across the membrane. Second, the membrane must be differentially permeable to different ions. If the membrane were entirely impermeable to ions, then no movement of ions across the membrane could occur, and no membrane potential would arise. If membranes are differentially permeable to ions, an electrical potential can arise across the membrane. Remember, synthetic bilayers are quite impermeable to ions, given the hydrophobicity of the internal part of the bilayer. Likewise, it is quite impermeable to glucose. It turns out that glial cells appear to have only a non-gated potassium channel, which allows the outward flow of potassium ions down the concentration gradient. The inside will then have a net negative charge since impermeable anions remain.

The chemical potential gradient causes this outward flow of potassium ions. As more ions leave, the inside becomes more negative, and a transmembrane potential develops that resists further potassium efflux. Eventually, they reach equilibrium, and the net efflux of potassium ceases. The resting transmembrane potential is around -75 mV, as we will derive below. Since glial cells appear only to express a non-gated (or leakage) potassium channel, their resting potential equals the potassium equilibrium potential. Figure \(\PageIndex{3}\) shows hypothetical transmembrane potentials (Ψ), and the electrical and chemical potentials (ΔG) for K⁺ loaded vesicles with and without a non-gated K⁺ channel

Figure \(\PageIndex{3}\): Visualizing the transmembrane potential in K⁺ loaded vesicles with and without a non-gated K⁺ channel

As shown previously, very little K⁺ efflux is required to develop a transmembrane potential. At equilibrium, the K⁺_IN is simply shown as < 0.1 M while K⁺_OUT is shown as > 0.1 M

A PhET simulation illustrates the differences between non-gated (leakage; always open) and gated channels (opening in response to stimuli such as ligands, transmembrane potential, or mechanical forces).

Neurotransmitter Activation of Neurons

What happens when a neurotransmitter binds to a receptor on the postsynaptic cell? We will study two examples. The first is the simplest: binding the neurotransmitter acetylcholine, released by a motor neuron, to its receptor on the muscle. This region is called the neuromuscular junction. Acetylcholine binding will lead to a transient depolarization of the muscle cell. Next, we will discuss how a neurotransmitter interacts with a postsynaptic neuron in the central nervous system. This is a much more complex system. Their differences are described below:

In neurons interacting with muscles:

• Most muscle fibers are innervated by only one neuron, a motor neuron
• Neurotransmitter release at the neuromuscular junction leads only to muscle excitation, not inhibition.
• All fibers are excited by the same neurotransmitter, acetylcholine.

In the central nervous system, life is more complicated:

• Stimuli are received from hundreds to thousands of different neurons.
• Nerves receive both excitatory and inhibitory stimuli from neurotransmitters.
• Different types of receptors detect stimuli and control the activity of various channels.
• Neuronal ion channels are gated by various mechanisms in addition to changes in membrane potential, including gating by heat, cold, stretch, or covalent modification.
• Most nerve cells have a resting potential of about -65 mV compared to -90 mV for a muscle cell.

What happens when a neurotransmitter binds to the receptor on the postsynaptic cell? A depolarization occurs (mediated by conformational changes in the transmitter-receptor complex), raising the membrane potential from the resting equilibrium level. What happens next depends on the identity of the post-synaptic cell. In the muscle cell, the rising potential caused by acetylcholine binding ultimately leads to muscle contraction by opening calcium channels in the membrane of the intracellular organelle. In a neuron, the rising potential triggers an action potential by opening voltage-gated sodium channels. The potential rises to about +35 mV but does not reach the Na⁺ equilibrium potential because the high positive potential opens a voltage-gated potassium channel. The potential then falls until it reaches the K⁺ equilibrium potential, thereby hyperpolarizing the cells. It slowly then relaxes back to the resting potential of -60 mV. This wave of potential changes sweeps along the post-synaptic membrane and underlies the neuron's "firing". A plot of transmembrane voltage changes vs time for a typical action potential is shown in Figure \(\PageIndex{4}\).

Figure \(\PageIndex{4}\): Action potential of a neuron. Original by en:User:Chris 73, updated by en:User:Diberri, GNU Free Documentation License,

Figure \(\PageIndex{5}\) shows the actual changes in ion permeability in various phases of the action potential (replace figure, seek permission)

mine

Figure \(\PageIndex{5}\): Na⁺ and K⁺ permeability during the action potential

Figure \(\PageIndex{6}\) shows an animation of a neuron firing, showing all the key players. (produced by PhET, University of Colorado, Boulder).

Proteins of the Neural Synapse

We must now account for the rise and fall of the membrane potential to a variety of neurotransmitters, including the cholinergic transmitters (ex., acetylcholine), catecholamines (dopamine, epinephrine, norepinephrine), amino acid derivatives (ex., Glu, Asp, N-methyl-D-Asp, Gly, gamma-amino-butyric acid -GABA), and peptides (endorphins, enkephalins). We will consider five membrane proteins as shown below in Figure \(\PageIndex{7}\).

Figure \(\PageIndex{7}\): 5 key membrane proteins involved in the action potential

We have explored many of these in detail in Chapter 11.3: Diffusion Across a Membrane—Channels, so in the following section, we will focus on just one new one, the chloride channel.

Na⁺-K⁺-ATPase: It transports sodium and potassium ions against a concentration gradient using ATP as an energy source. The protein is a sodium-dependent ATPase. The membrane potential could not be maintained without this protein since the sodium and potassium gradients would collapse. It also contributes to the potential since it is an electrogenic antiporter. (We have also seen that ungated potassium and sodium channels are present.)

Neurotransmitter receptor: The receptors we will consider here are typically ligand-gated ion channels. Once the ligand binds, the protein undergoes a conformational change, allowing ions to flow down their concentration gradient. Depending on the nature of the ion, the channel either initiates depolarization (when Na⁺ enters from the outside, raising ΔΨ) or inhibits depolarization (when Cl^- enters from the outside, lowering ΔΨ). When chloride channels open, they hyperpolarize the transmembrane potential. Stimulatory neurotransmitters (like glutamate) lead to membrane depolarization, while inhibitory neurotransmitters (like gamma-aminobutyric acid) lead to membrane hyperpolarization (making the membrane potential more negative).

Na⁺ channel (voltage-gated): When the membrane depolarizes to threshold, voltage-gated sodium channels undergo a conformational change and open, allowing Na⁺ ions to flow into the cell, thereby raising the membrane potential to a positive value. 33 mV (a value lower than the equilibrium sodium potential). This membrane protein is a voltage-gated channel, not a ligand-gated one.

Two potent neurotoxins, tetrodotoxin (from Pufferfish) and saxitoxin, bind to the channel and act as antagonists (inhibit the activity of the receptor by blocking sodium influx). Their structures are shown in Figure \(\PageIndex{8}\).

Figure \(\PageIndex{8}\): Structures of tetrodotoxin and saxitoxin

The guanidino group of tetrodotoxin appears to bind to the channel's entrance with high affinity, interacting with the hydrated sodium ion. Affinity chromatography using tetrodotoxin beads has been used to purify the protein. Figure \(\PageIndex{9}\) shows the relative sizes of ions used to probe size requirements for the channel. The hydrated K⁺ ion can not pass through.

Figure \(\PageIndex{9}\): Na⁺ channel ion selectivity

Depolarization of the membrane potential may cause outward movement and rotation of positively charged helices containing Lys and Arg side chains, presumably forming salt bridges (ion-ion interactions) with negatively charged side chains within the protein. Depolarization of the membrane disrupts a few salt bridges and effectively causes the movement of 1 or 2 charges along the helix through the membrane. Work occurs when charges are moved through an electric field. Work is also related to the ΔG of the system, which depends on the ratio of the open and closed forms of the channel. Other voltage-gated ion channels (for potassium and chloride) have a similar membrane topology and an S4 voltage-sensor helix.

K⁺ channel (voltage-gated): When the membrane potential reaches around +25 mV, the K⁺ channel, a voltage-gated membrane protein, changes conformation, allowing K⁺ efflux from the cell. This lowers the potential until it reaches the potassium equilibrium potential. It then slowly relaxes back to the cell resting potential of about -60 mV.

Cl^- channel: If these channels (typically ligand-gated) are open, they will hyperpolarize the cell and make it more difficult to fire.

The selectivity filter comprises many stacked rings of oxygens that can interact with a dehydrated K ion but not with a dehydrated Na⁺ ion, which can not approach close enough to form significant interactions. Surrounding the filter are twelve aromatic amino acids that constrain the pore opening size. The interactions of the filter O's with the K ion make up for the energetically disfavored dehydration of the ion. The filter contains two K⁺ ions that repel each other, thereby facilitating the vectorial discharge of the ions through the membrane. These ions must form weak interactions with the selectivity filter. The actual pore is primarily hydrophobic, which facilitates ion flow. The central cavity of the pore can accommodate water molecules in addition to K ions, thereby stabilizing them within the pore.

Inhibitory Neurotransmitters - The GABA Receptor

The main inhibitory neurotransmitters are GABA (gamma-aminobutyric acid), made from glutamic acid through decarboxylation of the α-C group, and glycine. They bind to transmitter-gated chloride channels, which, when open, hyperpolarize (make more negative) the transmembrane potential. Benzodiazepines (like Valium and Librium - anti-anxiety and muscle-relaxing agents) and barbiturates (like phenobarbital hypnotics) bind at allosteric sites on the GABA receptor and potentiate the binding of each other and GABA. This receptor is also affected by alcohol and anesthetics. Let's focus on the GABAA channels (also called the GABA(A) receptor or GABA(A)R) since they bind so many interesting pharmaceutical drugs.

Figure \(\PageIndex{10}\) shows an interactive iCn3D model of the human full-length heteromeric α1-β3-γ2L GABA(A)R in complex with picrotoxin, GABA, and megabody Mb38 (6huj) derived from cryo-EM (loads slowly).

Figure \(\PageIndex{10}\): Human full-length heteromeric alpha1beta3gamma2L GABA(A)R in complex with picrotoxin, GABA and megabody Mb38 (6huj) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...VdExv4QhFPsVX8 (loads slowly)

This structure is a Type-A γ-aminobutyric acid (GABA _A ) receptor. It consists of two alpha chains (orange), two beta chains (magenta), and one gamma chain (brown). Two GABAs (spacefill, CPK colors) are bound between the orange and magenta subunits (between the α and β chains). Picrotoxin is shown in spacefill CPK colors in the central pore (bottom), near the cytoplasmic end (blue bilayer dummy atoms). The extracellular domain (above the red) has glycans shown in SNFG cartoon form and shown in Figure \(\PageIndex{11}\).

Figure \(\PageIndex{11}\): Glycan structure for the Type-A γ-aminobutyric (GABA _A ) receptor

The structure of the receptor bound to many pharmacological agents has been solved. The structure of GABA (the agonist), bicuculline (a competitive antagonist), the benzodiazepines alprazolam (Valium) and diazepam (Xanax), a channel blocker (picrotoxin), ethanol, and Ro-15-4513 (an ethanol antagonist) are shown in Figure \(\PageIndex{12}\).

(\PageIndex{12}\): Structures of GABA-R binding molecules -GABA (the agonist), bicuculline (a competitive antagonist), the benzodiazepines alprazolam (Valium) and diazepam (Xanax), a channel blocker (picrotoxin), ethanol, and Ro-15-4513 (an ethanol antagonist)

The benzodiazepine diazepam (Xanax) binds at an allosteric site and promotes GABA binding, so it is considered an "indirect agonist”. It is also an anxiolytic (a drug used to reduce anxiety) and an anticonvulsant. Ethanol activates the inhibitory GABA channel. Too much ethanol leads to hyperpolarized cells, so neural responses are greatly inhibited, possibly leading to death. Ethanol acts synergistically with benzodiazepines, which makes this combination so lethal.

Figure \(\PageIndex{13}\) shows an interactive iCn3D model of the human full-length heteromeric α1-β3-γ2L GABA(A)R in complex with diazepam (Valium), GABA (6HUP) (loads slowly). https://structure.ncbi.nlm.nih.gov/i...fTZMqePnpdwTy8

Figure \(\PageIndex{13}\): Human full-length heteromeric α1-β3-γ2L GABA(A)R in complex with diazepam (Valium), GABA (6HUP). (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...fTZMqePnpdwTy8

The structure shows two alpha chains (orange), two beta chains (magenta), and one gamma chain (gray). Two GABAs (spacefill, CPK colors) are bound between the orange and magenta subunits (between the α and β chains). Three Valiums (spacefill, CPK) are shown in the structure. One is bound to an allosteric site between the orange (alpha) and gray (gamma) chains. Two others are bound within the bilayer, this time between the alpha (orange) and beta chains (magenta). Different benzodiazepines appear to bind in slightly different sites, and binding elicits their specific effects.

A drug, Ro-15-4513, was developed in the 1980s that antagonizes the effect of ethanol. It has a complicated pharmacology. It is considered a benzodiazepine “partial inverse agonist”. It has no effect by itself. It reverses the anticonvulsant effects of benzodiazepines and blocks the Cl^- effects of ethanol, so it is an inverse agonist of GABA-mediated Cl^- flux. Hence, it antagonizes the effect of benzodiazepines and alcohol. If given to intoxicated mice, they act normally! Such drugs have not reached the market as their use poses significant ethical issues.

The effect of pharmacological agents on the GABA receptor

In the “voltmeters” below, draw an arrow indicating if the transmembrane potential becomes more negative or more positive for the conditions given.

Answer

Add texts here. Do not delete this text first.

Metabotropic Neural Receptors

Some signaling molecules, whose effects are regulated by kinases (β-adrenergic and some olfactory signals by PKA, and acetylcholine by PKC, for example), are neurotransmitters. In all the previous examples, neurotransmitters directly gate inactive ion channels. These types of membrane receptors are classified as ionotropic. Typical examples of neurotransmitter-gated ion channels are the acetylcholine receptor in neuromuscular junctions and the central nervous system's Glu, Gly, and GABA receptors. These receptors are multimeric proteins. Receptors that directly gate ion flow are fast, with activities lasting milliseconds, and are used to elicit quick behavioral responses.

However, ion channels can also be indirectly gated when a neurotransmitter binds to its receptor, triggering downstream events that open an ion channel distinct from the receptor. In this case, the occupied receptor indirectly communicates to an ion channel through a G protein, for example. Examples of this indirect gating of ion channels include the serotonin, adrenergic, and dopamine receptors in the brain. These receptors are classic single-protein, serpentine GPCRs with seven transmembrane helices and intracellular domains that can interact with G proteins, as described above, to increase second-messenger levels (cAMP, DAG) in the neuron. The receptors are classified as metabotropic because they must activate a series of metabolic steps before ion channels open. The second messengers can either activate kinases in the cell, which phosphorylate ion channels to open or close them, or bind directly to the channel and modulate its activity through an allosteric conformational change. In some cases, the G protein directly acts on the ion channel. These different ways are described in Figure \(\PageIndex{14}\).

Figure \(\PageIndex{14}\): Neurotransmission: Gating through G Linked Receptors

In contrast to direct gating, receptors that indirectly gate ion channels exhibit slow activity lasting seconds to minutes. These receptors are usually involved in modulating behavior by altering neuronal excitability and the strength of neural connections, thereby influencing learning and memory. These changes can occur in many ways, summarized below:

Phosphorylating ion channels: Receptors that act through second-messenger systems can modulate ion channel activity by activating kinases that phosphorylate the channels. This may:

• open the channel normally closed at the resting potential, producing an effect like gating.
• Close a channel that usually opens at the resting potential (such as non-gated K channels, which, when closed, would depolarize the cell and make it more excitable).

Gα interaction with ion channels:

• the Ga subunit of the G protein interacts with K channels after stimulation of the CNS Acetylcholine receptor, opening the channel and hyperpolarizing the cell

Second messenger interaction with ion channels:

• cGMP opens cation channels in retinal cells after activation of the photoreceptor by photons
• cAMP opens cation channels in olfactory cells after activation of the olfactory receptor by odorants.

Second messenger effects on proteins other than ion channels (usually different receptors):

• the β-adrenergic receptors are phosphorylated by PKA and PKC (activated by stimulation of a different neurotransmitter receptor linked through a G protein to produce increased levels of second messengers cAMP and diacylglycerol). When phosphorylated, the β-adrenergic receptor, itself activated through G protein) can't bind Gs. This attenuates the β-adrenergic receptor's response to its neurotransmitter, leading to desensitization to that signal.

Second messengers regulate gene expression:

• cAMP-activated PKA can phosphorylate an inactive transcription factor, which can then bind to a DNA sequence called the cAMP Response Element (CRE) upstream of specific genes, leading to their transcription. The transcription factor is called CREB for cAMP Response Element Binding protein. Example: Tyrosine hydroxylase (a monooxygenase) is a key enzyme in the synthesis of epinephrine and norepinephrine. The activity of this protein is increased when PKA phosphorylates it. Hence, this modification of the already present protein can quickly increase its activity. If an animal is subjected to severe or long-term stress (e.g., cold or immobilization), presynaptic norepinephrine cells are stimulated to release the neurotransmitter. This requires continual synthesis of the neurotransmitter by the presynaptic cell. The increase in the synthesis of this neurotransmitter is caused by stimulation of the presynaptic cell by another neuron, leading to increased cAMP levels and, ultimately, activation of CREB, which increases transcription of the hydroxylase gene.

Caffeine

Caffeine produces a state of arousal in the central nervous system. High levels block the binding of an inhibitory neurotransmitter, adenosine, to the A2A adenosine receptor. Without caffeine, adenosine levels rise during the day, promoting its interaction with its receptor and leading to increased sleepiness and reduced concentration. When adenosine binds normally to its receptors, it activates the adenylate cyclase cascade, which activates PKA, leading to changes in the phosphorylation state of many proteins inside the cell, including protein phosphatase 2A. These changes inhibit neural firing. Caffeine blocks these changes.

Hallucinogenic drugs

Illicit drugs like LSD, psilocybin, and ecstasy can produce hallucinations as they have profound effects on consciousness and perception of self and reality. Recent clinical studies have shown that under tightly controlled conditions and doses, these drugs might have significant therapeutic effects in the treatment of mental health issues such as depression and post-traumatic stress disorder. Hence, their mechanisms of action have been the source of many studies.

All these drugs bind to the human serotonin 2A receptor (5-HT_2AR), a metabotropic GPCR receptor of serotonin (5-hydroxytryptamine). When serotonin binds to its receptor, it activates GPCR signaling, partly through beta-arrestins. These adapter proteins form complexes with ligand-bound and activated GPCRs, as well as with GPCR protein kinases.

Structures show that LSD binds to the orthosteric binding site (i.e., the active site, not an allosteric site) for serotonin. The study also found that serotonin and psilocin binding extends into an adjacent site called the extended binding pocket (EBP). These differences suggest that it should be possible to design agonists that have therapeutic, but not hallucinogenic, properties. Serotonin binding does not cause hallucinations. A drug, IHCH-7086, in mouse studies does not seem to provoke hallucinations but appears to have antidepressant effects (based on observations of mouse behaviors like twitching and freezing). The structures of serotonin (5HT) and psychotropic drugs that bind the 5HT_2AR are shown in Figure \(\PageIndex{15}\).

Figure \(\PageIndex{15}\): Structures of serotonin (5HT) and psychotropic drugs that bind the 5HT_2AR

Figure \(\PageIndex{16}\) shows a model of the serotonin receptor (5HT-2A) with each of the bound drugs. Note the different occupancy of the orthosteric and extended binding pockets.

Figure \(\PageIndex{16}\): The serotonin receptor (5HT-2A) with bound serotonin and orthosteric binding analogs.

The GPCRs have been aligned for each of the poses. The ligands are represented in spacefill in a single color, as listed below:

• red: serotonin (5HT), the physiological agonist, 7WC4
• yellow: psilocin, 7WC5 LSD, 7WC6
• magenta: LSD, 7WC6
• cyan: IHCH-7086

An alternative explanation for why serotonin does not cause hallucinations and why the other agonists do is that, in addition to binding plasma membrane GPCRs, the agonist might have additional effects that are associated with hallucinations by binding intracellular membrane GPCRs. 

The cerebral cortex plays a key role in what it means to be human.  It is key to higher-level processing involving thought, learning, emotion, personality, language, and memory.  As such, its dysregulation is involved in mental health conditions such as depression, et al., in which synaptic plasticity is decreased as evidenced by a reduction in dendritic structure required for synaptic connections.  The serotonin (5HT) reuptake inhibitors (SSRIs) used to treat depression appear to increase neural plasticity and connections over time (so they don't give immediate responses).  Hallucinogens that target the 5-HT2A receptor (5-HT2AR) also promote neuroplasticity.  However, neuronal growth in cortical cultures is unaffected by serotonin.  What is the mechanism that allows various combinations of neuron plasticity and hallucinations, all from the same receptor?

One factor determining these differential effects is that serotonin (5-HT) is more polar and cannot readily cross cell membranes. At the same time, the HT2AR agonists that are hallucinogenic are less polar (more lipophilic) and could potentially enter cells where they might elicit hallucinogenic effects.  Intracellular hallucinogens would bind internal 5-HT2AR receptors, which are found in cortical neurons on the Golgi and other organelles that are more acidic than the cytoplasm or extracellular environment.  (In fact, in cortical neurons, the main location of the 5-HT2ARs is intracellular.) This might lead to more prolonged retention of intracellular hallucinogens and longer signaling (LSD has a profound hallucinogenic effect that lasts for 10 hours or more), leading to neuronal growth as well.

If serotonin is given to wild-type mice, no hallucinations occur, as evidenced by the lack of a head twitch response (HTR).  However, when given to mutant mice expressing serotonin transporter (SERT) on cortical neurons, the HTR response was observed. The import of serotonin led to neuroplasticity.  It could be that serotonin is not the physiological ligand for intracellular HT2ARs.  It might also imply that, as with the case of endocannabinoids, we have endogenous psychedelics.  The subtle difference in the binding of serotonin and the hallucinogens in Figure 16 might have little to do with their tendency to produce hallucinations.

Recent Updates:  LSD binding to the BDNF receptor TrkB   06/09/23

Much is still not known about how antidepressants work and what causes delays in their therapeutic effects.  Increased synaptic connections through expanded neuroplasticity appear to be required for their therapeutic effects.  The primary action of drugs like Prozac, a serotonin reuptake inhibitor, is to block the serotonin transporter (SERT), thereby increasing extracellular serotonin levels at the neural synapse.  Tricyclic antidepressants as well as monoamine oxidase inhibitors (MAOI), also increase monoamine neurotransmitters in the synapse with delayed therapeutic effects (the monoamines act quickly, however).  Perhaps these agents bind other receptors!

They do.  The binding of both typical and fast antidepressants also occurs in the transmembrane domain of tyrosine kinase receptor 2 (TRKB), the brain-derived neurotrophic factor (BDNF) receptor, which is associated with increased neuroplasticity. The receptor also binds cholesterol, which modulates its activity.  The antidepressant binding site is formed on dimerization of their transmembrane domains.  Mutations in the transmembrane region block the efficacy of the antidepressants. 

LSD and other psychedelics also produce fast and long-lasting antidepressant effects promoted by increases in neuroplasticity.  Studies have shown that LSD and psilocin bind to slightly overlapping sites in the transmembrane domain of the BDNF receptor as antidepressants, but with a 1000-fold higher affinity.  If the LSD binding site on the serotonin 2A receptor (5-HT_2A) is blocked, LSD still has antidepressant and increased neuroplasticity effects.  

Figure \(\PageIndex{17}\) shows the interaction of LSD and the metabolite of psilocybin, psilocin (PSI), with the TRKB receptor. 

Figure \(\PageIndex{17}\): Characterization of the psychedelics binding site in the TrkB TMD.  Moliner, R., Girych, M., Brunello, C.A. et al. Psychedelics promote plasticity by directly binding to the BDNF receptor TrkB. Nat Neurosci 26, 1032–1041 (2023). https://doi.org/10.1038/s41593-023-01316-5.  Creative Commons Attribution 4.0 International License. http://creativecommons.org/licenses/by/4.0/.

Panel a–c show representative MD snapshots showing the binding pocket for LSD (purple) (a) and PSI (green) (c) in the extracellular-facing crevice of the TrkB TMD dimer (gray). Side chains (yellow) of relevant binding site residues are displayed. A structural model of full-length TrkB dimer (gray) embedded in a lipid membrane is shown with bound BDNF (blue) and LSD (purple) (b). 

Panel d shows in silico binding free energy estimations for fluoxetine, LSD, and PSI. Each free energy estimate (ΔG, circles) and its statistical error (SE, bars) were estimated from a separate set of FEP simulations (n = 1). Dissociation constants are given as a range with upper and lower bounds converted from ΔG − SE and ΔG + SE, respectively. 

Panels e and f show chemical structures of LSD (e) and PSI (f) with atom numbers annotated. 

Panel i shows the distributions of TMD dimer C-terminal distance and that LSD and PSI stabilize the cross-shaped conformation of TrkB favorable for receptor activation in a 40 mol% CHOL membrane. Lines represent the mean distribution, and bands represent the standard errors (n = 10 independent simulations). TMD conformations corresponding to indicated C-terminal distances and drug-bound states are shown in the inset. 

Figure \(\PageIndex{18}\) shows the different binding modes of LSD and Fluoxetine (Prozac), a selective serotonin reuptake inhibitor (SSRI) for TrkB.  Fluoxetine is used to treat depression, obsessive-compulsive disorder (OCD), bulimia nervosa, and panic disorder.

 Figure \(\PageIndex{18}\): Different TrkB binding modes of LSD and fluoxetine. Moliner et al., ibid.

Panels a,b, show representative snapshots of atomistic MD simulations showing the front (a) and back (b) views of the binding pockets for LSD (purple) and fluoxetine (yellow) in the extracellular-facing crevice of TrkB TMD dimers. Side chains of relevant binding site residues are displayed. Superimposed structures of TrkB optimally bound to LSD or fluoxetine reveal that, although some residues involved in binding are shared (Y433 and V437), the binding modes differ. Fluoxetine binds at a site deeper within the dimer, locking the TMD dimers in a more open cross-shaped conformation (distance between the center of mass L451–L453 Cα atoms of each monomer ~20 Å). In contrast, LSD binds closer to the N-terminus of the TrkB TMD and establishes more stable interactions with the dimer: a hydrogen bond between the oxygen atom of the diethylamide group of LSD and the Y433 residue of one monomer, and pi-stacking of the aromatic backbone of the drug with the Y433 residue of the second monomer, locking the TMD dimer in a tighter cross-shaped conformation (L451–L453 Cα distance ~17 Å) compared with fluoxetine. Drugs are shown in van der Waals representation. Oxygen, nitrogen, and hydrogen atoms are shown in red, blue, and white, respectively.

Summary

(Summary written by Claude, Sonnet 4.6, Anthropic)

Neural signaling depends on the precise control of ion gradients and transmembrane potentials across neuronal membranes. The Na⁺-K⁺-ATPase establishes the resting electrochemical gradient by exporting three Na⁺ ions for every two K⁺ ions imported, creating a resting membrane potential of approximately -65 mV in neurons. This potential is maintained by the differential permeability of the membrane to ions through constitutively open (non-gated) and stimulus-responsive (gated) channels. The selectivity of ion channels arises not from simple size exclusion but from a narrow selectivity filter where ions are partially dehydrated and form specific, transient interactions with carbonyl or hydroxyl oxygen atoms lining the pore.

An action potential is initiated when a neurotransmitter binds to a postsynaptic ligand-gated receptor, depolarizing the membrane above threshold. This opens voltage-gated Na⁺ channels, causing rapid Na⁺ influx and further depolarization to approximately +35 mV. The subsequent opening of voltage-gated K⁺ channels drives K⁺ efflux, repolarizing and transiently hyperpolarizing the membrane before it returns to its resting potential. These events propagate as a wave down the axon. Neurotransmission at the neuromuscular junction is simpler—acetylcholine acts on a single excitatory receptor type—while central nervous system neurons integrate thousands of excitatory and inhibitory inputs using diverse receptor classes.

The GABA_A receptor, the principal inhibitory ionotropic receptor, is a pentameric Cl⁻ channel whose opening hyperpolarizes the membrane and reduces neuronal excitability. Its multiple pharmacological binding sites—orthosteric GABA sites at α-β subunit interfaces, benzodiazepine allosteric sites at α-γ interfaces, and a channel-blocking site for picrotoxin—make it a target for anxiolytics, anticonvulsants, anesthetics, and alcohol. Benzodiazepines potentiate GABA binding as positive allosteric modulators, explaining both their therapeutic utility and their dangerous synergy with ethanol, which also activates the receptor through a distinct mechanism.

Metabotropic receptors—GPCRs for serotonin, dopamine, norepinephrine, adenosine, and acetylcholine—modulate neuronal excitability more slowly by engaging second-messenger cascades that phosphorylate ion channels, regulate transcription via CREB, or promote receptor desensitization. Caffeine exemplifies this by blocking the inhibitory adenosine A2A receptor and thereby maintaining higher cAMP levels and neural firing rates. Psychedelic drugs, including LSD and psilocybin, bind the serotonin 5-HT2A receptor, and emerging evidence suggests their hallucinogenic effects may involve access to intracellular GPCRs on Golgi membranes in cortical neurons—a site inaccessible to the more polar endogenous agonist serotonin. Most strikingly, both classical antidepressants (SSRIs such as fluoxetine) and psychedelics promote neuroplasticity through a shared but pharmacologically distinct binding site in the transmembrane domain dimer of TrkB, the BDNF receptor, with LSD binding approximately 1000-fold more tightly than fluoxetine through hydrogen bonding and π-stacking interactions with Tyr433 that stabilize a tighter cross-shaped TMD dimer conformation. These findings illuminate why psychedelics may offer faster and more durable antidepressant effects than conventional medications.