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Transduction of Biologic Signals

Receptor Theory
Definition

In the broadest sense, a receptor is a component of a cell that interacts selectively with an extracellular compound to initiate a cascade of biochemical events that culminate in the observed effects of the compound. Binding of a drug to a receptor determines (1) the quantitative relationship between a given dose of a drug and the resulting effect, (2) the selectivity of a given drug's activity and effect, and (3) an explanation of the pharmacologic activity of receptor agonists, antagonists, and inverse agonists. Receptors therefore mediate or amplify the effect of a drug on the biologic system.

Historical Perspective

Although the overall concept of receptors is generally attributed to Paul Ehrlich (1854–1915), earlier work performed by Claude Bernard (1813–1878) paved the way for receptor theory. This earlier work is of particular interest to anesthesiologists because it elucidated the mechanism of action of curare, an arrow poison. In his experiments, Claude Bernard ligated vessels leading to one hind limb of a frog while leaving nerve input intact; he then administered intravenous curare. Pinching the paralyzed hind limb produced reflex movements in the opposite unparalyzed vessel-ligated hind limb. These experiments and others demonstrated for the first time the separation between sensory and motor nervous systems and revealed that circulating "substances" produced selective effects on organ systems, a concept important in the development of receptor theory.

The concept of selectivity of effect eventually led Paul Ehrlich to his conclusion that "agents cannot act unless they are bound," a cornerstone of receptor theory. Although Ehrlich is attributed with discovering this concept, his contemporary J. N. Langley (1852–1926) coined the term receptive substance (i.e., receptor). Langley extended Claude Bernard's work by showing that nicotine and curare had mutually antagonistic effects on the same receptive substance, which was neither nerve nor muscle. We now recognize this receptive substance as the nicotinic acetylcholine receptor in the neuromuscular junction. Several decades of research have refined the receptor theory, and receptors are now recognized as discrete excitable proteins.

Classic Receptor Theory

The binding of a ligand (i.e., drug molecule) to its receptor follows the laws of chemical reactions (analogous to Michaelis Menten enzyme kinetics) and can be summarized by the following relationship:





In this reversible equation, kon is the rate constant for the ligand binding to the receptor, koff is the rate constant for the ligand disassociating from the receptor, [L] is the concentration of the ligand, [R] is the concentration of the unbound receptor, and [LR] is the concentration of the bound receptor. Units for kon are units of [L] multiplied by the units of [R] over time; typically, units of L are nanomoles per liter. The units for koff are the units of [LR] over time. The term Kd , the dissociation constant, mathematically defines characteristics of ligand-receptor interactions at equilibrium:





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The units of Kd are the units of [L], which are units of concentration. When enough drug is administered to occupy exactly 50% of the receptors (i.e., [R] = [LR]), the measured Kd value is equal to the drug concentration. A low Kd value indicates that less drug is required to occupy 50% of the receptors, implying that each molecule of drug is tightly associated with the receptor. A high Kd value indicates that more drug is required to occupy 50% of receptors, implying a weak binding to the receptor. Kd has been determined for many of the drugs administered during anesthesia. The reciprocal of Kd is Ka , the association constant, which is exactly analogous to the Ka defined for protein binding. Ka is a measure of the affinity of the drug for the receptor. A drug with a low Kd value has a high Ka value and therefore high affinity for the receptor.

In practice, it is difficult to measure precise drug-receptor occupancy. It is often assumed that the drug-receptor complex represents an intermediate step in the production of a specific effect. Many investigators apply pharmacodynamic theory to compare a given dose of a drug with a resulting effect. An effect can be any biochemical or physiologic variable that is measurable. A measured effect can be an alteration in a biochemical compound, an enzyme level, a physiologic variable such as heart rate or blood pressure, or a response to any graded input into the biologic system. For example, in evaluating the pharmacodynamics of muscle relaxants, the measured effect is not a direct measurement of drug-receptor complexes, but rather a response to a neuromuscular stimulus as delivered by a nerve stimulator.

From the discussion of pharmacokinetics, the delivery of a drug to a given receptor is clearly time and dose dependent. If the receptor is located within the central compartment, there may be nearly instantaneous equilibration after an intravenous injection, and the peak effect may occur immediately. If drug must cross from the central compartment to the site of drug effect, a delay will occur. The onset of drug effect can also be delayed by the time required for the body to respond to the drug. This delay may occur anywhere on the path between drug-receptor binding and clinical manifestation of drug effect. Examples of post-transduction time delays are drug-receptor complex-induced secondary messenger changes, increased enzyme synthesis, and the time required for physiologic change (e.g., reduced fluid content).

RECEPTOR AGONISTS AND ANTAGONISTS.

Drugs that are agonists induce an effect that mimics endogenous hormones or neurotransmitters when bound to a receptor. This effect may be stimulatory or inhibitory. The term affinity as related to a given agonist is a measure of the attraction between the given drug and receptor. A drug with low affinity for a given receptor tends not to bind to the receptor and produces little or no effect. An agonist that binds avidly (or with high affinity) to a given receptor produces the receptor-determined effect at a lower given dose.

Full agonists completely activate a receptor, whereas partial agonists only partially activate a receptor, even at very high concentrations ( Fig. 3-23 ). The difference between full and partial agonists results from different intrinsic efficacy for each drug. Efficacy should not be confused with affinity. Two drugs can have identical affinity for a receptor (and therefore bind to the same extent at a given


Figure 3-23 Effects of various types of ligands on receptor responses. A full agonist produces complete (100%) activation of a receptor at high concentrations, whereas partial agonist binding results in less than 100% activation, even at very high concentrations. A neutral antagonist has no activity of its own. Inverse agonists can be thought of as "superantagonists" because binding of these ligands produces a response below the baseline response measured in the absence of drug. If the physiologic effect of the baseline levels of activated receptor (R*) is small, antagonists and inverse agonists may not be clinically distinguishable.

drug concentration), but they may produce different levels of activation of the receptor. Both are agonists, but the drug that produces maximal receptor activation is called a full agonist and has high efficacy, whereas the drug that produces less than a maximal receptor activation is called a partial agonist with less intrinsic efficacy. The mechanisms underlying one drug's producing full versus partial agonist effects remain shrouded in mystery, but they may be related to the ability of the drug to stabilize the high-affinity state of the receptor (see "Receptor States").

Drugs that are antagonists inhibit or prevent receptor-mediated agonist effects by competing for receptor occupancy. A competitive antagonist can generally be displaced from the receptor complex by the administration of a large enough concentration of receptor agonist, permitting the agonist to produce the expected effect despite the presence of antagonist. A noncompetitive antagonist usually binds irreversibly to the receptor complex, producing loss of the expected effect that cannot be compensated by administration of high concentrations of receptor agonist. For example, vecuronium is a competitive antagonist of acetylcholine (see Chapter 13 ). Acetylcholine mediates muscle contraction through the postsynaptic nicotinic acetylcholine receptor at the neuromuscular junction. When vecuronium binds to the postsynaptic acetylcholine receptor, acetylcholine agonism is blocked, and neuromuscular transmission is inhibited. The result is flaccid paralysis. Neuromuscular transmission may be reinstated by administering an acetylcholine esterase inhibitor. Acetylcholine esterase inhibitors prevent breakdown of acetylcholine, effectively raising the concentration of the

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agonist, acetylcholine, at the receptor, which then displaces vecuronium from the receptor complex. When enough vecuronium is displaced, muscle contraction is reinstated. Most antagonist drugs used in clinical medicine are competitive antagonists.

Inverse agonists can be thought of as "superantagonists" because they decrease receptor responses to less than baseline values (see Fig. 3-23 ). An example of concentration-response relationships for four different benzodiazepines and EEG response[10] is shown in Figure 3-24 (see Chapter 10 ). In this example, the EEG response represents a surrogate clinical end point for binding of benzodiazepines to the γ-aminobutyric acid (GABAA ) ligand-gated ion channel complex with resultant modulations of ion fluxes in the central nervous system (CNS).

RECEPTOR STATES.

Classic receptor theory describes interaction between ligand and receptor based on the laws of mass action. At a molecular level, this interaction has been interpreted over the years to suggest that binding of agonist to receptor initiates a change in receptor conformation, changing the receptor from an inactive (R) to an


Figure 3-24 The concentration-electroencephalographic (EEG) response relationship for four benzodiazepines: midazolam (full agonist, a), bretazenil (partial agonist, b), flumazenil (antagonist, c), and R0 19-4063 (inverse agonist, d). The maximum effect seen in the EEG response correlates with clinical action (full agonist > partial agonist > antagonist > inverse agonist). (Adapted from Shafer S: Principles of pharmacokinetics and pharmacodynamics. In Longnecker DE, Tinker JH, Morgan GE [eds]: Principles and Practice of Anesthesiology, 2nd ed. St. Louis, Mosby-Year Book, 1997, and from Mandema JW, Kuck MT, Danhof M: Differences in intrinsic efficacy of benzodiazepines are reflected in their concentration-EEG effect relationship. Br J Pharmacol 105:164–170, 1992.)

activated (R*) state ( Fig. 3-25 ). In this model, the change in receptor conformation facilitates activated receptor coupling to intermediary proteins (e.g., guanine nucleotide [G] proteins) or second messengers (i.e., effectors [E]), which then initiate a rapid cascade of cellular responses.

Transgenic animal experiments have shed light on ligand and receptor interactions. In an elegant set of experiments in which β2 -adrenergic receptors were massively overexpressed in mouse myocardium, second messenger responses were increased in the absence of ligand when compared with normal mouse myocardium. Activity of adenylyl cyclase, the second messenger most commonly studied for β2 -adrenergic receptors, is identical at baseline in transgenic animals (in the absence of drug) compared with normal mice stimulated with isoproterenol, a β2 -adrenergic-receptor agonist. Isoproterenol stimulation of transgenic animals is unable to increase adenylyl cyclase activity higher than this elevated baseline. This finding suggests that a subpopulation of receptors is already fully activated in these transgenic animals. One way to analyze these findings is to consider that at baseline a small

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Figure 3-25 Schematic drawing of classic receptor activation. For years, binding of ligand was thought to cause receptors to change from an inactive state (R) to an activated state (R*). The activated receptor interacted with intermediary guanine nucleotide (G) proteins or directly with second messenger cascades (i.e., effectors [E]), or both. Later information suggests a more complicated equilibrium between R and R*, as shown in Figure 2-26 and Figure 2-27.

percentage (e.g., 1%) of receptors spontaneously exists in the activated (R*) state. Because this 1% of activated receptors may be normal in that tissue, the activity of these R* receptors is included as part of the baseline effect measured in the absence of drug. However, after massive overexpression of β2 -adrenergic receptors occurs (e.g., in a transgenic animal), 1% of the total receptors becomes a very large number of activated receptors. This mimics the effect of a maximal dose of drug, which would produce similar numbers of receptors in the R* state.

These findings have broad implications for understanding receptor states and drug-receptor interactions. Instead of ligand binding causing a change from R to R* (see Fig. 3-25 ), these data suggest spontaneous conversion from the R to the R* state ( Fig. 3-26 ). In most tissues, the concentration of R* represents a small fraction of total overall receptor (hence the large arrow pointing toward R in Fig. 3-26 ). However, baseline receptor responses are


Figure 3-26 Spontaneous conversion of receptors from an inactive (R) to an active (R*) state. In most tissues, R* represents a small fraction of the total receptor population.

known to vary among tissues, a finding suggesting that the absolute concentration of R* (and the spontaneous conversion rates between R and R*) may be tissue dependent. This finding is clinically important because inverse agonists, which are known to stabilize R, drive the equation toward the left (i.e., toward R). This decreases responses to less than the clinical baseline value, toward the true baseline in the absence of any receptor activation. The biggest differences between apparent baseline and true baseline are likely to occur in tissues with higher concentrations of spontaneous R*, and inverse agonists are therefore most efficacious in such tissues. Figure 3-24 illustrates a tissue (i.e., CNS) with enough activated GABAA ligand-gated ion channels (R*) to see EEG changes with an inverse agonist. This finding provides a rationale for the inverse agonist antiepileptic drugs being developed. This discussion assumes only two receptor states, R (completely inactive) and R* (fully activated). Later experimental evidence suggests the existence of intermediate transition states between R and R* (hence the use of curved lines in addition to straight lines in Fig. 3-26 ). However, in the interest of simplicity, further discussion in this chapter uses only R and R*.

The discovery of spontaneous conversion between R and R* has implications for understanding the action of ligands (i.e., agonists, antagonists, and inverse agonists). Instead of agonists causing conversion from R to R*, it is thought that ligands stabilize a given receptor state. As shown in Figure 3-27 , agonists stabilize (or energetically favor) R*, and inverse agonists stabilize R, whereas neutral antagonists bind equally to R and R*. Binding of an agonist to a receptor removes the receptor from the equation because rapid coupling to second messengers occurs, driving (by mass action) the equation toward that specific state ( Fig. 3-28 ). The probability of being in the R* state increases in the presence of an agonist. Conversely, inverse agonists drive the equation to the left, stabilizing the R configuration. Neutral antagonists bind equally to R and R*, preventing agonist binding without altering the equilibrium between R and R*; neutral antagonists do not change the observed baseline response (see Fig. 3-23 ). Many classic antagonists have been tested and found to be inverse agonists rather than neutral antagonists. For example, among the commonly clinically used β2 -adrenergic-receptor (β2 AR) antagonists, metoprolol and bisoprolol show significant inverse agonist activity, carvedilol has weaker inverse agonist properties at the β2 AR and displays


Figure 3-27 Ligand stabilization of receptor states. Agonists stabilize active receptors (R*), inverse agonists stabilize inactive receptors (R), and neutral antagonists bind equally to R and R* without affecting the equilibrium between the two receptor states.

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Figure 3-28 Equilibrium between inactive receptors (R) and active receptors (R*) is tissue specific and depends on the type of ligand administered. By stabilizing R*, agonists drive the equilibrium to the right. Inverse agonists stabilize R, driving the equilibrium to the left. Because neutral antagonists bind equally to R and R*, they do not affect tissue-specific equilibrium between R and R*.

neutral antagonist activity at β1 ARs, alprenolol has weaker still (20%) inverse agonist activity at the β2 AR, and xamoterol displays marked partial agonist activity at β1 ARs.[11] [12] Because inverse agonists may have theoretical advantages in treating certain diseases (e.g., seizures, congestive heart failure), clinicians should critically evaluate recent pharmacologic data before using an antagonist to distinguish whether the drug is a neutral antagonist or an inverse agonist.

Receptor Structure
General Principles

Receptors are located in many places in the cell—outer cell membrane, cytoplasm, intracellular organelle membranes, and nucleus. The overall physical structure of a


Figure 3-29 Schematic drawing of four types of membrane-associated drug targets: membrane (G protein-coupled) receptor, ligand-gated ion channel, voltage-sensitive ion channel, and an enzyme. Potential sites of drug action are shown. βAR, β-adrenergic receptor; GABAA , γ-aminobutyric acid.

receptor depends on the type of receptor considered and its location. For example, membrane receptors have structures different from those of cytoplasmic receptors. In general, cytoplasmic receptors and nuclear receptors bind ligands that readily traverse cell-membrane lipid bilayers to reach the cytoplasm or nucleus. These ligands therefore must contain significant hydrophobic (i.e., lipophilic) components. Examples of cytoplasmic and nuclear receptors include steroid and thyroid hormone receptors. In contrast, most receptors important to the anesthesiologist are excitable cell-membrane proteins, such as membrane receptors, ligand-gated ion channels, and voltage-gated ion channels ( Fig. 3-29 ). Many endogenous hormones and exogenous drugs are water soluble (i.e., hydrophilic and charged) and are unable to cross cell-membrane lipid bilayers. A mechanism is required to transduce drug binding at the extracellular surface into changes in intracellular physiology. Overall, the membrane's receptor structure depends on whether coupling to second messengers through intermediary proteins is required or whether the receptor is part of a larger ion channel complex (e.g., ligand-gated ion channels). Each of these types of membrane receptors has a role in mediating the action of various drugs important in anesthesiology. It is beyond the scope of this chapter to list all identified receptor proteins. However, prototypical examples are used to illustrate important pharmacodynamic principles.

Guanine Nucleotide Protein-Coupled Receptors

One of the most common methods by which membrane receptors translate agonist occupancy into intracellular action is through intermediary G proteins. As a result, G protein-coupled receptors are the most abundant type of receptor known. The overall structure of a G protein-coupled receptor is shown schematically in Figure 3-30 . Two-dimensional representation reveals an extracellular amino terminus containing glycosylation sites, an intracellular carboxyl terminus, a fatty acid attachment (usually

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Figure 3-30 Schematic drawing of a G protein-coupled receptor. A two-dimensional version of receptor structure is shown with seven transmembrane domains, an extracellular amino (NH2 ) terminus (with associated glycosylation sites [Y]), an intracellular carboxyl (COOH) terminus, palmitoylated cysteine residue (crooked line extending into the membrane), three extracellular loops, and four intracellular loops. Major sites of G protein interactions with the receptor are speckled; potential sites of phosphorylation in the third intracellular loop and carboxyl terminus are enclosed in boxes.

through palmitoylation or myristoylation of a carboxylterminal cysteine residue), three extracellular loops, and four intracellular loops. Major sites of G protein interactions with the receptor occur in the third and fourth (and to a lesser extent, second) intracellular loops, whereas major sites of phosphorylation, which is often correlated with desensitization (i.e., dampening of receptor responses), generally occur in the third intracellular loop and carboxyl terminus.

Although a two-dimensional schematic is shown in Figure 3-30 , it is important to remember that G protein-coupled receptors are three-dimensional structures with transmembrane segments coalescing around a central binding pocket ( Fig. 3-31 ). [13] Although transmembrane amino acids must be hydrophobic overall to be energetically favored in the lipid membrane, side chains on individual amino acids may be charged. Specific amino acids therefore act as counter ions to anchor charged (water-soluble) hormone or drug to the receptor. Structure-activity relationships between drug and the three-dimensional configuration of the receptor's binding site are critically important in determining binding properties. The chemical structure of a drug must match the three-dimensional configuration of the binding area of a receptor. Subtle changes in drug structure may dramatically alter the ability of a drug to bind to a specific receptor population. Two drugs with seemingly unrelated two-dimensional chemical structures may bind to the same receptor if their three-dimensional structures and charged areas are similar. The three-dimensional components of a ligand that interact with ligand-recognition portions of a receptor are referred to as the pharmacophore.


Figure 3-31 Schematic drawing of the three-dimensional structure of the β2 -adrenergic receptor (a prototypical G protein-coupled receptor), looking from the outside of the cell inward. Transmembrane domains (denoted by cylinders and numbered with roman numerals) coalesce to form a binding pocket. The correct orientation of the agonist norepinephrine is shown. Notice that agonist affinity is determined by specific amino acids located in transmembranes III, IV, and VII. These critical sites in transmembrane domains have been determined experimentally using chimeric receptor and mutagenesis approaches combined with sophisticated computer-modeling techniques. Ultimate confirmation of these structures will require crystallographic data, an approach that is being attempted by several laboratories. (Adapted from Stapelfeldt WH: The autonomic nervous system. In Schwinn DA [ed]: Scientific Principles of Anesthesia, vol 2. Philadelphia, Current Medicine, 1998.)

Ion Channels

In addition to membrane receptors, pharmacologic agents act on other excitable cell-membrane proteins, such as ion channels. These channels mediate neural signaling by modulating ion permeability in electrically excitable membranes. Because many drugs act directly or indirectly through ion channels, it is important for the anesthesiologist to understand their general function. Classic ion channels such as the sodium channel have charged regions that span the cell membrane. The formation of ion pairs between many positive and negative charges helps to stabilize the channel in the membrane. Voltage-dependent gating of the sodium channel is made possible by the presence of a voltage sensor—a collection of charges that moves under the influence of the cell-membrane electrical field (hence the name voltage-gated ion channels). During depolarization, these charges presumably move outward, causing conformational changes and rearrangement of ion pairs that result in sodium permeability. Local anesthetics work by blocking voltage-gated sodium channels. This concept is discussed in more detail in Chapter 14 .

The combination of a classic receptor protein plus an ion channel results in a ligand-gated ion channel, giving rise to the unique ability of certain drugs to alter membrane permeability to ions directly. Many anesthetic drugs act on ligand-gated ion channels, such as the nicotinic acetylcholine receptor and the GABAA receptor. The structure of the GABAA receptor is shown in Figure 3-32 .[14] [15] Ligand-gated ion channels have hydrophobic and charged

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Figure 3-32 Cross-section of a schematized γ-aminobutyric acid (GABA)-benzodiazepine receptor complex, which is a prototypical ligand-gated ion channel complex. Binding of benzodiazepine agonists to the GABAA ligand-gated ion channel facilitates the action of endogenous GABA. This results in increased inhibitory chloride ion flux in the central nervous system. A, Schematic drawing of the GABAA ligand-gated ion channel complex shows the receptor sites for benzodiazepines and GABA, as well as distinct receptor sites for barbiturates and alcohol. B, Model of the GABAA receptor and chloride ion channel shows that the protein complex is composed of a hetero-oligomer of five subunits, including α, β, γ, and δ or ρ polypeptides. Each subunit has four putative membrane-spanning domains (numbered 1 through 4 and represented by cylinders). (A, from Berkowitz DE: Cellular signal transduction. In Schwinn DA [ed]: Scientific Principles of Anesthesia, vol 2. Philadelphia, Current Medicine, 1998; B, adapted from Firestone L, Quinlan J, Gyulai F: Mechanisms of anesthetic action. In Schwinn DA [ed]: Scientific Principles of Anesthesia, vol 2. Philadelphia, Current Medicine, 1998.)

transmembrane regions. The hydrophobic regions stabilize the structure in the membrane, and the centrally charged region serves as the pore for ion flux. The binding of drugs to receptors in ligand-gated ion channel complexes usually enhances or dampens an already existing neurotransmitter-induced ion flux. For example, the neurotransmitter GABA binds to its receptor within the GABAA ligand-gated ion channel complex and causes the movement of chloride ions intracellularly. This action results in lowered membrane potential, a hallmark of inhibitory neurotransmission. Drugs (e.g., benzodiazepines, barbiturates, alcohols) binding to other receptor sites in the GABAA ligand-gated ion channel facilitate the action of endogenous GABA. This process results in increased inhibitory chloride ion
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flux in the CNS (see Fig. 3-32 ). Most hypnotics (i.e., benzodiazepines, barbiturates, propofol, etomidate, and possibly the hypnotic action of inhaled anesthetics) act through potentiation of endogenous GABA at the GABAA ligand gated ion channel.

Ion Pumps

Another type of excitable membrane protein is the ion pump. The sodium-potassium-adenosine triphosphatase (ATPase) pump is perhaps the most familiar ion pump to the anesthesiologist because it is inhibited by digitalis. Extracellular fluid has high sodium and low potassium levels, whereas intracellular fluid has high potassium and low sodium concentrations. Because the nerve at rest is selectively permeable to potassium, but not to sodium, potassium moves extracellularly, creating a net positive extracellular charge and a net negative intracellular charge. Action potentials activate sodium channels, allowing sodium to rush intracellularly along chemical and electrical gradients. The sodium-potassium-ATPase pump then rapidly pumps sodium out of the cell in exchange for potassium, returning the cell to its original cation composition and electrical gradient. Drugs that act on ion pumps alter intracellular-extracellular cation ratios, resulting in altered membrane electrical potential.

Digitalis acts by inhibiting the sodium-potassium-ATPase pump. This is of special importance in the myocardial cell, where sodium-potassium-ATPase exchange is replaced by slower sodium-calcium exchange, thereby increasing intracellular calcium concentrations. Because calcium increases myocardial contractility, improved myocardial pump function results. This is the basis of digitalis use in the treatment of congestive heart failure.

Second Messengers

The binding of a hormone or drug to its receptor does not instantly produce clinical effects. Instead, a series of rapid biochemical events couples receptor binding to ultimate clinical effects. These biochemical events are called second messengers. Because alterations in second messenger coupling can alter the effectiveness of a drug, it is important that the anesthesiologist understand the general principles of second messenger action.

Many membrane receptors couple to their primary second messenger through G proteins, which are intermediate regulatory molecules. The coupling of G proteins to receptor-hormone complexes requires energy in the form of guanosine triphosphate (GTP). The hydrolysis of G protein-associated GTP to GDP is regulated by another set of proteins, called RGS proteins. After the receptor interacts with the G protein, the biochemical reaction in the effector cascade is triggered. There are stimulating (e.g., Gs , Gq ) and inhibitory (e.g., Gi , Go ) G proteins, and the physiologic effect is determined by the specific G protein and subsequent cellular response. G proteins are heterotrimeric, composed of three subunits: α, β, and γ. Receptor and G protein interactions result in dissociation of the G protein subunits into α and βγ. The α-subunit of most G proteins confers specificity between receptor and effectors. Although βγ-subunits were originally thought simply to anchor G proteins to the cell membrane, dissociated βγ-subunits are clearly capable of directly stimulating second messengers. The βγ-subunits play a role in anchoring regulatory kinases to the cell membrane, leading to enhanced phosphorylation of membrane receptors.

One of the best understood second messenger systems is the adenylyl cyclase system. In this system, complexes of stimulatory G protein receptors and hormones increase activity of the enzyme adenylyl cyclase, resulting in increased levels of cyclic adenosine 3',5'-monophosphate (cAMP) in the cell ( Fig. 3-33 ). Inhibitory G protein-receptor-hormone complexes decrease the activity of adenylyl cyclase, resulting in decreased levels of cellular cAMP. In general, increased levels of intracellular cAMP activate protein kinases that phosphorylate various proteins, ion channels, and second messenger enzymes. For example, β-adrenergic-receptor stimulation results in stimulation of adenylyl cyclase activity, increasing production of intracellular cAMP, which activates protein kinase A, which phosphorylates the L-type calcium channel in myocardium to produce increases in intracellular calcium, ultimately leading to increased inotropy. Examples of drugs or hormones that activate adenylyl cyclase include glucagon and histamine. In contrast, muscarinic (M2 , M4 ) agonists, α2 -adrenergic agonists, and adenosine generally inhibit adenylyl cyclase.

Another example of a second messenger system is the phosphatidylinositol system. Hydrolysis of phosphatidyl-inositol-4,5-bisphosphate (PIP2 ) in the cell membrane, catalyzed by Gq activation of phospholipase C, generates two main second messenger molecules, inositol-1,4,5-trisphosphate (IP3 ) and 1,2-diacylglycerol (DAG). IP3 then mobilizes intracellular calcium from nonmitochondrial intracellular stores by interacting with distinct IP3 receptors on the surface of these organelles. The resulting increase in intracellular calcium levels produces biologic effects such as smooth muscle contraction. Examples of drugs that mediate phosphatidylinositol hydrolysis include α1 -adrenergic, endothelin, and muscarinic (M1 , M3 , M5 ) agonists.

Stimulation of receptors and second messengers ultimately leads to physiologic effects in a given tissue. The physiologic effects produced depend on the presence of


Figure 3-33 Schematic drawing of the signal transduction cascade of the β-adrenergic receptor (βAR). ATP, adenosine triphosphate; βγ, stimulatory G protein βγ subunit; cAMP, cyclic adenosine monophosphate; Gs α, α-subunit of the stimulatory G protein (Gs ).

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Figure 3-34 Adrenergic receptor subtype physiology. HR, heart rate; NE, norepinephrine.

specific receptor subtypes, G proteins, and second messengers within that tissue. An example of the wide-ranging cardiovascular effects produced by a single hormone—norepinephrine, the predominant sympathetic neurotransmitter—can be seen in Figure 3-34 . Effects of sympathetic stimulation vary according to tissue. For example, acute effects of β2 -adrenergic-receptor-mediated vasodilation enhance skeletal muscle blood flow, β-adrenergic-receptor-mediated inotropic effects improve myocardial performance, and adrenergic effects in the CNS result in enhanced alertness; all of these effects enable effective performance during stressful situations. Chronic stimulation of stress responses, however, leads to other adrenergically mediated effects, some maladaptive, such as excessive myocardial hypertrophy, hypertension, and dampening of myocardial function.

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