Introduction to the Pharmacology of CNS Drugs: Introduction

Drugs acting in the central nervous system (CNS) were among the first to be discovered by primitive humans and are still the most widely used group of pharmacologic agents. In addition to their use in therapy, many drugs acting on the CNS are used without prescription to increase one's sense of well-being.

The mechanisms by which various drugs act in the CNS have not always been clearly understood. In the last three decades, however, dramatic advances have been made in the methodology of CNS pharmacology. It is now possible to study the action of a drug on individual cells and even single ion channels within synapses. The information obtained from such studies is the basis for several major developments in studies of the CNS.

First, it is clear that nearly all drugs with CNS effects act on specific receptors that modulate synaptic transmission. A very few agents such as general anesthetics and alcohol may have nonspecific actions on membranes (although these exceptions are not fully accepted), but even these non–receptor-mediated actions result in demonstrable alterations in synaptic transmission.

Second, drugs are among the most important tools for studying all aspects of CNS physiology, from the mechanism of convulsions to the laying down of long-term memory. As described below, agonists that mimic natural transmitters (and in many cases are more selective than the endogenous substances) and antagonists are extremely useful in such studies. Natural Toxins: Tools for Characterizing Ion Channels, describes a few of these substances.

Third, unraveling the actions of drugs with known clinical efficacy has led to some of the most fruitful hypotheses regarding the mechanisms of disease. For example, information on the action of antipsychotic drugs on dopamine receptors has provided the basis for important hypotheses regarding the pathophysiology of schizophrenia. Studies of the effects of a variety of agonists and antagonists on -aminobutyric acid (GABA) receptors has resulted in new concepts pertaining to the pathophysiology of several diseases, including anxiety and epilepsy.

This chapter provides an introduction to the functional organization of the CNS and its synaptic transmitters as a basis for understanding the actions of the drugs described in the following chapters.

Methods for the Study of CNS Pharmacology

Like many areas of science, major progress in the study of CNS drugs has depended on the development of new experimental techniques. The first detailed description of synaptic transmission was made possible by the invention of glass microelectrodes, which permit intracellular recording. The development of the brain slice technique permitted an analysis of the physiology and pharmacology of synapses. Detailed electrophysiologic studies of the action of drugs on both voltage- and transmitter-operated channels were further facilitated by the introduction of the patch clamp technique, which permits the recording of current through single channels. Channels can be expressed in cultured cells and the currents evoked by their activation recorded (Figure 21–1). Histochemical, immunologic, and radioisotopic methods have made it possible to map the distribution of specific transmitters, their associated enzyme systems, and their receptors. Molecular cloning has had a major impact on our understanding of CNS receptors. These techniques make it possible to determine the precise molecular structure of the receptors and their associated channels. Finally, mice with mutated genes for specific receptors or enzymes (knockout mice) can provide important information regarding the physiologic and pharmacologic roles of these components.

Ion Channels & Neurotransmitter Receptors

The membranes of nerve cells contain two types of channels defined on the basis of the mechanisms controlling their gating (opening and closing): voltage-gated and ligand-gated channels (Figure 21–2A and B). Voltage-gated channels respond to changes in the membrane potential of the cell. The voltage-gated sodium channel described in Chapter 14 for the heart is an example of the first type of channel. In nerve cells, these channels are concentrated on the initial segment and the axon and are responsible for the fast action potential, which transmits the signal from cell body to nerve terminal. There are many types of voltage-sensitive calcium and potassium channels on the cell body, dendrites, and initial segment, which act on a much slower time scale and modulate the rate at which the neuron discharges. For example, some types of potassium channels opened by depolarization of the cell result in slowing of further depolarization and act as a brake to limit further action potential discharge.

Neurotransmitters exert their effects on neurons by binding to two distinct classes of receptor. The first class is referred to as ligand-gated channels, or ionotropic receptors. The receptor consists of subunits, and binding of ligand directly opens the channel, which is an integral parts of the receptor complex (see Figure 22–6). These channels are insensitive or only weakly sensitive to membrane potential. Activation of these channels typically results in a brief (a few milliseconds to tens of milliseconds) opening of the channel. Ligand-gated channels are responsible for fast synaptic transmission typical of hierarchical pathways in the CNS (see following text).

The second class of neurotransmitter receptor is referred to as metabotropic receptors. These are 7-transmembrane G protein-coupled receptors of the type described in Chapter 2. The binding of neurotransmitter to this type of receptor does not result in the direct gating of a channel. Rather, binding to the receptor engages a G protein, which results in the production of second messengers that modulate voltage-gated channels. These interactions can occur entirely with the plane of the membrane and are referred to as membrane-delimited pathways (Figure 21–2C). In this case, the G protein (often the subunit) interacts directly with the voltage-gated ion channel. In general, two types of voltage-gated ion channels are the targets of this type of signaling: calcium channels and potassium channels. When G proteins interact with calcium channels, they inhibit channel function. This mechanism accounts for the presynaptic inhibition that occurs when presynaptic metabotropic receptors are activated. In contrast, when these receptors are postsynaptic, they activate (cause the opening of) potassium channels, resulting in a slow postsynaptic inhibition. Metabotropic receptors can also modulate voltage-gated channels less directly by the generation of diffusible second messengers (Figure 21–2D). A classic example of this type of action is provided by the adrenoceptor, which generates cAMP via the activation of adenylyl cyclase (see Chapter 2). Whereas membrane-delimited actions occur within microdomains in the membrane, second messenger-mediated effects can occur over considerable distances. Finally, an important consequence of the involvement of G proteins in receptor signaling is that, in contrast to the brief effect of ionotropic receptors, the effects of metabotropic receptor activation can last tens of seconds to minutes. Metabotropic receptor predominate in the diffuse neuronal systems in the CNS (see below).

The Synapse & Synaptic Potentials

The communication between neurons in the CNS occurs through chemical synapses in the majority of cases. (A few instances of electrical coupling between neurons have been documented, and such coupling may play a role in synchronizing neuronal discharge. However, it is unlikely that these electrical synapses are an important site of drug action.) The events involved in synaptic transmission can be summarized as follows.

An action potential in the presynaptic fiber propagates into the synaptic terminal and activates voltage-sensitive calcium channels in the membrane of the terminal (see Figure 6–3). The calcium channels responsible for the release of transmitter are generally resistant to the calcium channel-blocking agents discussed in Chapter 12 (verapamil, etc) but are sensitive to blockade by certain marine toxins and metal ions (see Tables 21–1 and 12–4). Calcium flows into the terminal, and the increase in intraterminal calcium concentration promotes the fusion of synaptic vesicles with the presynaptic membrane. The transmitter contained in the vesicles is released into the synaptic cleft and diffuses to the receptors on the postsynaptic membrane. Binding of the transmitter to its receptor causes a brief change in membrane conductance (permeability to ions) of the postsynaptic cell. The time delay from the arrival of the presynaptic action potential to the onset of the postsynaptic response is approximately 0.5 ms. Most of this delay is consumed by the release process, particularly the time required for calcium channels to open.

The first systematic analysis of synaptic potentials in the CNS was in the early 1950s by Eccles and associates, who recorded intracellularly from spinal motor neurons. When a microelectrode enters a cell, there is a sudden change in the potential recorded by the electrode, which is typically about –70 mV (Figure 21–3). This is the resting membrane potential of the neuron. Two types of pathways—excitatory and inhibitory—impinge on the motor neuron.

When an excitatory pathway is stimulated, a small depolarization or excitatory postsynaptic potential (EPSP) is recorded. This potential is due to the excitatory transmitter acting on an ionotropic receptor, causing an increase in cation permeability. Changing the stimulus intensity to the pathway, and therefore the number of presynaptic fibers activated, results in a graded change in the size of the depolarization. When a sufficient number of excitatory fibers are activated, the excitatory postsynaptic potential depolarizes the postsynaptic cell to threshold, and an all-or-none action potential is generated.

When an inhibitory pathway is stimulated, the postsynaptic membrane is hyperpolarized owing to the selective opening of Cl^– channels, producing an inhibitory postsynaptic potential (IPSP) (Figure 21–4). However, because the equilibrium potential for Cl^– is only slightly more negative than the resting potential (~ –65 mV), the hyperpolarization is small and contributes only modestly to the inhibitory action. The opening of the Cl^– channel during the inhibitory postsynaptic potential makes the neuron "leaky" so that changes in membrane potential are more difficult to achieve. This shunting effect decreases the change in membrane potential during the excitatory postsynaptic potential. As a result, an excitatory postsynaptic potential that evoked an action potential under resting conditions fails to evoke an action potential during the inhibitory postsynaptic potential (Figure 21–4). A second type of inhibition is presynaptic inhibition. It was first described for sensory fibers entering the spinal cord, where excitatory synaptic terminals receive synapses called axoaxonic synapses (described later). When activated, axoaxonic synapses reduce the amount of transmitter released from the terminals of sensory fibers. It is interesting that presynaptic inhibitory receptors are present on almost all presynaptic terminals in the brain even though axoaxonic synapses appear to be restricted to the spinal cord. In the brain, transmitter spills over to neighboring synapses to activate the presynaptic receptors.

Sites of Drug Action

Virtually all the drugs that act in the CNS produce their effects by modifying some step in chemical synaptic transmission. Figure 21–5 illustrates some of the steps that can be altered. These transmitter-dependent actions can be divided into presynaptic and postsynaptic categories.

Drugs acting on the synthesis, storage, metabolism, and release of neurotransmitters fall into the presynaptic category. Synaptic transmission can be depressed by blockade of transmitter synthesis or storage. For example, reserpine depletes monoamine synapses of transmitter by interfering with intracellular storage. Blockade of transmitter catabolism inside the nerve terminal can increase transmitter concentrations and has been reported to increase the amount of transmitter released per impulse. Drugs can also alter the release of transmitter. The stimulant amphetamine induces the release of catecholamines from adrenergic synapses (see Chapters 6 and 32). Capsaicin causes the release of the peptide substance P from sensory neurons, and tetanus toxin blocks the release of transmitters. After a transmitter has been released into the synaptic cleft, its action is terminated either by uptake or by degradation. For most neurotransmitters, there are uptake mechanisms into the synaptic terminal and also into surrounding neuroglia. Cocaine, for example, blocks the uptake of catecholamines at adrenergic synapses and thus potentiates the action of these amines. However, acetylcholine is inactivated by enzymatic degradation, not reuptake. Anticholinesterases block the degradation of acetylcholine and thereby prolong its action. No uptake mechanism has been found for any of the numerous CNS peptides, and it has yet to be demonstrated whether specific enzymatic degradation terminates the action of peptide transmitters.

In the postsynaptic region, the transmitter receptor provides the primary site of drug action. Drugs can act either as neurotransmitter agonists, such as the opioids, which mimic the action of enkephalin, or they can block receptor function. Receptor antagonism is a common mechanism of action for CNS drugs. An example is strychnine's blockade of the receptor for the inhibitory transmitter glycine. This block, which underlies strychnine's convulsant action, illustrates how the blockade of inhibitory processes results in excitation. Drugs can also act directly on the ion channel of ionotropic receptors. For example, barbiturates can enter and block the channel of many excitatory ionotropic receptors. In the case of metabotropic receptors, drugs can act at any of the steps downstream of the receptor. Perhaps the best example is provided by the methylxanthines, which can modify neurotransmitter responses mediated through the second-messenger cAMP. At high concentrations, the methylxanthines elevate the level of cAMP by blocking its metabolism and thereby prolong its action.

The traditional view of the synapse is that it functions like a valve, transmitting information in one direction. However, it is now clear that the synapse can generate signals that feed back onto the presynaptic terminal to modify transmitter release. Endocannabinoids are the best documented example of such retrograde signaling. Postsynaptic activity leads to the synthesis and release of endocannabinoids, which then bind to receptors on the presynaptic terminal. Although the gas nitric oxide (NO) has long been proposed as a retrograde messenger, its physiologic role in the CNS is still not well understood.

The selectivity of CNS drug action is based almost entirely on the fact that different transmitters are used by different groups of neurons. Furthermore, these transmitters are often segregated into neuronal systems that subserve broadly different CNS functions. Without such segregation, it would be impossible to selectively modify CNS function, even if one had a drug that operated on a single neurotransmitter system. That such segregation does occur has provided neuroscientists with a powerful pharmacologic approach for analyzing CNS function and treating pathologic conditions.

Identification of Central Neurotransmitters

Because drug selectivity is based on the fact that different pathways use different transmitters, a primary goal of neuropharmacologists is to identify the transmitters in CNS pathways. Establishing that a chemical substance is a transmitter has been far more difficult for central synapses than for peripheral synapses. The following criteria have been established for transmitter identification.

Localization

Approaches that have been used to prove that a suspected transmitter resides in the presynaptic terminal of the pathway under study include biochemical analysis of regional concentrations of suspected transmitters and immunocytochemical techniques for enzymes and peptides.

Release

To determine whether the substance is released from a particular region, local collection (in vivo) of the extracellular fluid can sometimes be accomplished. In addition, slices of brain tissue can be electrically or chemically stimulated in vitro and the released substances measured. To determine whether the release is relevant to synaptic transmission, it is important to establish that the release is calcium-dependent.

Synaptic Mimicry

Finally, application of the suspected substance should produce a response that mimics the action of the transmitter released by nerve stimulation. Furthermore, application of a selective antagonist should block the response. Microiontophoresis, which permits highly localized drug administration, has been a valuable technique in assessing the action of suspected transmitters. Because of the complexity of the CNS, specific pharmacologic antagonism of a synaptic response provides a particularly powerful technique for transmitter identification.

Cellular Organization of the Brain

Most of the neuronal systems in the CNS can be divided into two broad categories: hierarchical systems and nonspecific or diffuse neuronal systems.

Hierarchical Systems

Hierarchical systems include all the pathways directly involved in sensory perception and motor control. The pathways are generally clearly delineated, being composed of large myelinated fibers that can often conduct action potentials at a rate of more than 50 m/s. The information is typically phasic and occurs in bursts of action potentials. In sensory systems, the information is processed sequentially by successive integrations at each relay nucleus on its way to the cortex. A lesion at any link incapacitates the system. Within each nucleus and in the cortex, there are two types of cells: relay or projection neurons and local circuitneurons (Figure 21–6A). The projection neurons that form the interconnecting pathways transmit signals over long distances. The cell bodies are relatively large, and their axons emit collaterals that arborize extensively in the vicinity of the neuron. These neurons are excitatory, and their synaptic influences, which involve ionotropic receptors, are very short-lived.

The excitatory transmitter released from these cells is, in most instances, glutamate. Local circuit neurons are typically smaller than projection neurons, and their axons arborize in the immediate vicinity of the cell body. Most of these neurons are inhibitory, and they release either GABA or glycine. They synapse primarily on the cell body of the projection neurons but can also synapse on the dendrites of projection neurons as well as with each other. Two common types of pathways for these neurons (Figure 21–6A) include recurrent feedback pathways and feed-forward pathways. A special class of local circuit neurons in the spinal cord forms axoaxonic synapses on the terminals of sensory axons (Figure 21–6B). In some sensory pathways such as the retina and olfactory bulb, local circuit neurons may actually lack an axon and release neurotransmitter from dendritic synapses in a graded fashion in the absence of action potentials.

Although there is a great variety of synaptic connections in these hierarchical systems, the fact that a limited number of transmitters are used by these neurons indicates that any major pharmacologic manipulation of this system will have a profound effect on the overall excitability of the CNS. For instance, selectively blocking GABA_A receptors with a drug such as picrotoxin results in generalized convulsions. Thus, although the mechanism of action of picrotoxin is specific in blocking the effects of GABA, the overall functional effect appears to be quite nonspecific, because GABA-mediated synaptic inhibition is so widely utilized in the brain.

Nonspecific or Diffuse Neuronal Systems

Neuronal systems that contain one of the monoamines—norepinephrine, dopamine, or 5-hydroxytryptamine (serotonin)—provide examples in this category. Certain other pathways emanating from the reticular formation and possibly some peptide-containing pathways also fall into this category. These systems differ in fundamental ways from the hierarchical systems, and the noradrenergic systems serve to illustrate the differences.

Noradrenergic cell bodies are found primarily in a compact cell group called the locus caeruleus located in the caudal pontine central gray matter. The number of neurons in this cell group is small, approximately 1500 on each side of the brain in the rat.

Because these axons are fine and unmyelinated, they conduct very slowly, at about 0.5 m/s. The axons branch repeatedly and are extraordinarily divergent. Branches from the same neuron can innervate several functionally different parts of the CNS. In the neocortex, these fibers have a tangential organization and therefore can monosynaptically influence large areas of cortex. The pattern of innervation by noradrenergic fibers in the cortex and nuclei of the hierarchical systems is diffuse, and these fibers form a very small percentage of the total number in the area. In addition, the axons are studded with periodic enlargements called varicosities, which contain large numbers of vesicles. In some instances, these varicosities do not form synaptic contacts, suggesting that norepinephrine may be released in a rather diffuse manner, as occurs with the noradrenergic autonomic innervation of smooth muscle. This indicates that the cellular targets of these systems are determined largely by the location of the receptors rather than by the location of the release sites. Finally, most neurotransmitters utilized by diffuse neuronal systems, including norepinephrine, act—perhaps exclusively—on metabotropic receptors and therefore initiate long-lasting synaptic effects. Based on these observations, it is clear that the monoamine systems cannot be conveying topographically specific types of information; rather, vast areas of the CNS must be affected simultaneously and in a rather uniform way. It is not surprising, then, that these systems have been implicated in such global functions as sleeping and waking, attention, appetite, and emotional states.

Central Neurotransmitters

A vast number of small molecules have been isolated from the brain, and studies using a variety of approaches suggest that the agents listed in Table 21–2 are neurotransmitters. A brief summary of the evidence for some of these compounds follows.

Amino Acids

The amino acids of primary interest to the pharmacologist fall into two categories: the acidic amino acid glutamate and the neutral amino acids glycine and GABA. All these compounds are present in high concentrations in the CNS and are extremely potent modifiers of neuronal excitability.

Glutamate

Excitatory synaptic transmission is mediated by glutamate, which is present in very high concentrations in excitatory synaptic vesicles (~100 mM). Glutamate is released into the synaptic cleft by Ca²⁺-dependent exocytosis (Figure 21–7). The released glutamate acts on postsynaptic glutamate receptors and is cleared by glutamate transporters present on surrounding glia. In glia, glutamate is converted to glutamine by glutamine synthetase, released from the glia, taken up by the nerve terminal, and converted back to glutamate by the enzyme glutaminase. The high concentration of glutamate in synaptic vesicles is achieved by the vesicular glutamate transporter (VGLUT).

Almost all neurons that have been tested are strongly excited by glutamate. This excitation is caused by the activation of both ionotropic and metabotropic receptors, which have been extensively characterized by molecular cloning. The ionotropic receptors can be further divided into three subtypes based on the action of selective agonists: -amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA),  kainic acid (KA)  and N -methyl-D -aspartate (NMDA). All the ionotropic receptors are composed of four subunits. AMPA receptors, which are present on all neurons, are heterotetramers assembled from four subunits (GluA1-GluA4). The majority of AMPA receptors contain the GluA2 subunit and are permeable to Na⁺ and K⁺, but not to Ca²⁺. Some AMPA receptors, typically present on inhibitory interneurons, lack the GluA2 subunit and are also permeable to Ca²⁺.

Kainate receptors are not as uniformly distributed as AMPA receptors, being expressed at high levels in the hippocampus, cerebellum, and spinal cord. They are formed from a number of subunit combinations (GluK1-GluK5). Although GluK4 and GluK5 are unable to form channels on their own, their presence in the receptor changes the receptor's affinity and kinetics. Similar to AMPA receptors, kainate receptors are permeable to Na⁺ and K⁺ and in some subunit combinations can also be permeable to Ca²⁺.

NMDA receptors are as ubiquitous as AMPA receptors, being present on essentially all neurons in the CNS. All NMDA receptors require the presence of the subunit GluN1. The channel also contains one or two NR2 subunits (GluN2A-D). Unlike AMPA and kainate receptors, all NMDA receptors are highly permeable to Ca²⁺ as well as to Na⁺ and K⁺. NMDA receptor function is controlled in a number of intriguing ways. In addition to glutamate binding, the channel also requires the binding of glycine to a separate site. The physiologic role of glycine binding is unclear because the glycine site appears to be saturated at normal ambient levels of glycine. Another key difference between AMPA and kainate receptors on the one hand, and NMDA receptors on the other, is that AMPA and kainate receptor activation results in channel opening at resting membrane potential, whereas NMDA receptor activation does not. This is due to the voltage-dependent block of the NMDA pore by extracellular Mg²⁺. When the neuron is strongly depolarized, as occurs with intense activation of the synapse or by activation of neighboring synapses, Mg²⁺ is expelled and the channel opens. Thus, there are two requirements for NMDA receptor channel opening: Glutamate must bind the receptor and the membrane must be depolarized. The rise in intracellular Ca²⁺ that accompanies channel opening results in a long-lasting enhancement in synaptic strength that is referred to as long-term potentiation (LTP). The change can last for many hours or even days and is generally accepted as an important cellular mechanism underlying learning and memory.

The metabotropic glutamate receptors are G protein-coupled receptors that act indirectly on ion channels via G proteins. Metabotropic receptors (mGluR1-mGluR8) have been divided into three groups (I, II, and III). A variety of agonists and antagonists have been developed that interact selectively with the different groups. Group I receptors are typically located postsynaptically and are thought to cause neuronal excitation by activating a nonselective cation channel. These receptors also activate phospholipase C, leading to IP₃-mediated intracellular Ca²⁺ release. In contrast, group II and group III receptors are typically located on presynaptic nerve terminals and act as inhibitory autoreceptors. Activation of these receptors causes the inhibition of Ca²⁺ channels, resulting in inhibition of transmitter release. These receptors are activated only when the concentration of glutamate rises to high levels during repetitive stimulation of the synapse. Activation of these receptors causes the inhibition of adenylyl cyclase and decreases cAMP generation.

The postsynaptic membrane at excitatory synapses is thickened and referred to as the postsynaptic density (PSD; Figure 21–7). This is a highly complex structure containing glutamate receptors, signaling proteins, scaffolding proteins, and cytoskeletal proteins. A typical excitatory synapse contains AMPA receptors, which tend to be located toward the periphery, and NMDA receptors, which are concentrated in the center. Kainate receptors are present at a subset of excitatory synapses, but their exact location is unknown. Metabotropic glutamate receptors (group I), which are localized just outside the postsynaptic density, are also present at some excitatory synapses.

GABA and Glycine

Both GABA and glycine are inhibitory neurotransmitters, which are typically released from local interneurons. Interneurons that release glycine are restricted to the spinal cord and brain stem, whereas interneurons releasing GABA are present throughout the CNS, including the spinal cord. It is interesting that some interneurons in the spinal cord can release both GABA and glycine. Glycine receptors are pentameric structures that are selectively permeable to Cl^–. Strychnine, which is a potent spinal cord convulsant and has been used in some rat poisons, selectively blocks glycine receptors.

GABA receptors are divided into two main types: GABA_A and GABA_B. inhibitory postsynaptic potentials in many areas of the brain have a fast and slow component. The fast component is mediated by GABA_A receptors and the slow component by GABA_B receptors. The difference in kinetics stems from the differences in coupling of the receptors to ion channels. GABA_A receptors are ionotropic receptors and, like glycine receptors, are pentameric structures that are selectively permeable to Cl^–. These receptors are selectively inhibited by picrotoxin and bicuculline, both of which cause generalized convulsions. A great many subunits for GABA_A receptors have been cloned; this accounts for the large diversity in the pharmacology of GABA_A receptors, making them key targets for clinically useful agents (see Chapter 22). GABA_B receptors are metabotropic receptors that are selectively activated by the antispastic drug baclofen. These receptors are coupled to G proteins that, depending on their cellular location, either inhibit Ca²⁺ channels or activate K⁺ channels. The GABA_B component of the inhibitory postsynaptic potential is due to a selective increase in K⁺ conductance. This inhibitory postsynaptic potential is long-lasting and slow because the coupling of receptor activation to K⁺ channel opening is indirect and delayed. GABA_B receptors are localized to the perisynaptic region and thus require the spillover of GABA from the synaptic cleft. GABA_B receptors are also present on the axon terminals of many excitatory and inhibitory synapses. In this case, GABA spills over onto these presynaptic GABA_B receptors, inhibiting transmitter release by inhibiting Ca²⁺ channels. In addition to their coupling to ion channels, GABA_B receptors also inhibit adenylyl cyclase and decrease cAMP generation.

Acetylcholine

Acetylcholine was the first compound to be identified pharmacologically as a transmitter in the CNS. Eccles showed in the early 1950s that excitation of Renshaw cells by motor axon collaterals in the spinal cord was blocked by nicotinic antagonists. Furthermore, Renshaw cells were extremely sensitive to nicotinic agonists. These experiments were remarkable for two reasons. First, this early success at identifying a transmitter for a central synapse was followed by disappointment because it remained the sole central synapse for which the transmitter was known until the late 1960s, when comparable data became available for GABA and glycine. Second, the motor axon collateral synapse remains one of the best-documented examples of a cholinergic nicotinic synapse in the mammalian CNS, despite the rather widespread distribution of nicotinic receptors as defined by in situ hybridization studies. Most CNS responses to acetylcholine are mediated by a large family of G protein-coupled muscarinic receptors. At a few sites, acetylcholine causes slow inhibition of the neuron by activating the M₂ subtype of receptor, which opens potassium channels. A far more widespread muscarinic action in response to acetylcholine is a slow excitation that in some cases is mediated by M₁ receptors. These muscarinic effects are much slower than either nicotinic effects on Renshaw cells or the effect of amino acids. Furthermore, this M₁ muscarinic excitation is unusual in that acetylcholine produces it by decreasing the membrane permeability to potassium, ie, the opposite of conventional transmitter action.

A number of pathways contain acetylcholine, including neurons in the neostriatum, the medial septal nucleus, and the reticular formation. Cholinergic pathways appear to play an important role in cognitive functions, especially memory. Presenile dementia of the Alzheimer type is reportedly associated with a profound loss of cholinergic neurons. However, the specificity of this loss has been questioned because the levels of other putative transmitters, eg, somatostatin, are also decreased.

Monoamines

Monoamines include the catecholamines (dopamine and norepinephrine) and 5-hydroxytryptamine. Although these compounds are present in very small amounts in the CNS, they can be localized using extremely sensitive histochemical methods. These pathways are the site of action of many drugs; for example, the CNS stimulants cocaine and amphetamine appear to act primarily at catecholamine synapses. Cocaine blocks the reuptake of dopamine and norepinephrine, whereas amphetamines cause presynaptic terminals to release these transmitters.

Dopamine

The major pathways containing dopamine are the projection linking the substantia nigra to the neostriatum and the projection linking the ventral tegmental region to limbic structures, particularly the limbic cortex. The therapeutic action of the antiparkinsonism drug levodopa is associated with the former area (see Chapter 28), whereas the therapeutic action of the antipsychotic drugs is thought to be associated with the latter (see Chapter 29). Dopamine-containing neurons in the tuberobasal ventral hypothalamus play an important role in regulating hypothalamohypophysial function. Five dopamine receptors have been identified, and they fall into two categories: D₁-like (D₁ and D₅) and D₂-like (D₂, D3, D₄). All dopamine receptors are metabotropic. Dopamine generally exerts a slow inhibitory action on CNS neurons. This action has been best characterized on dopamine-containing substantia nigra neurons, where D₂-receptor activation opens potassium channels via the G_i coupling protein.

Norepinephrine

Most noradrenergic neurons are located in the locus caeruleus or the lateral tegmental area of the reticular formation. Although the density of fibers innervating various sites differs considerably, most regions of the CNS receive diffuse noradrenergic input. All noradrenergic receptor subtypes are metabotropic. When applied to neurons, norepinephrine can hyperpolarize them by increasing potassium conductance. This effect is mediated by ₂ receptors and has been characterized most thoroughly on locus caeruleus neurons. In many regions of the CNS, norepinephrine actually enhances excitatory inputs by both indirect and direct mechanisms. The indirect mechanism involves disinhibition; that is, inhibitory local circuit neurons are inhibited. The direct mechanism involves blockade of potassium conductances that slow neuronal discharge. Depending on the type of neuron, this effect is mediated by either ₁ or receptors. Facilitation of excitatory synaptic transmission is in accordance with many of the behavioral processes thought to involve noradrenergic pathways, eg, attention and arousal.

5-Hydroxytryptamine

Most 5-hydroxytryptamine (5-HT, serotonin) pathways originate from neurons in the raphe or midline regions of the pons and upper brain stem. 5-HT is contained in unmyelinated fibers that diffusely innervate most regions of the CNS, but the density of the innervation varies. 5-HT acts on more than a dozen receptor subtypes. Except for the 5-HT₃ receptor, all of these receptors are metabotropic. The ionotropic 5-HT₃ receptor exerts a rapid excitatory action at a very limited number of sites in the CNS. In most areas of the CNS, 5-HT has a strong inhibitory action. This action is mediated by 5-HT_1A receptors and is associated with membrane hyperpolarization caused by an increase in potassium conductance. It has been found that 5-HT_1A receptors and GABA_B receptors activate the same population of potassium channels. Some cell types are slowly excited by 5-HT owing to its blockade of potassium channels via 5-HT₂ or 5-HT₄ receptors. Both excitatory and inhibitory actions can occur on the same neuron. It has often been speculated that 5-HT pathways may be involved in the hallucinations induced by LSD (lysergic acid) , since this compound can antagonize the peripheral actions of 5-HT. However, LSD does not appear to be a 5-HT antagonist in the CNS, and typical LSD-induced behavior is still seen in animals after raphe nuclei are destroyed. Other proposed regulatory functions of 5-HT-containing neurons include sleep, temperature, appetite, and neuroendocrine control.

Peptides

A great many CNS peptides have been discovered that produce dramatic effects both on animal behavior and on the activity of individual neurons. Many of the peptides have been mapped with immunohistochemical techniques and include opioid peptides (eg, enkephalins, endorphins), neurotensin, substance P, somatostatin, cholecystokinin, vasoactive intestinal polypeptide, neuropeptide Y, and thyrotropin-releasing hormone. As in the peripheral autonomic nervous system, peptides often coexist with a conventional nonpeptide transmitter in the same neuron. A good example of the approaches used to define the role of these peptides in the CNS comes from studies on substance P and its association with sensory fibers. Substance P is contained in and released from small unmyelinated primary sensory neurons in the spinal cord and brain stem and causes a slow excitatory postsynaptic potential in target neurons. These sensory fibers are known to transmit noxious stimuli, and it is therefore surprising that—although substance P receptor antagonists can modify responses to certain types of pain—they do not block the response. Glutamate, which is released with substance P from these synapses, presumably plays an important role in transmitting pain stimuli. Substance P is certainly involved in many other functions because it is found in many areas of the CNS that are unrelated to pain pathways.

Many of these peptides are also found in peripheral structures, including peripheral synapses. They are described in Chapters 6 and 17.

Nitric Oxide

The CNS contains a substantial amount of nitric oxide synthase (NOS), which is found within certain classes of neurons. This neuronal NOS is an enzyme activated by calcium-calmodulin, and activation of NMDA receptors, which increases intracellular calcium, results in the generation of nitric oxide. Although a physiologic role for nitric oxide has been clearly established for vascular smooth muscle, its role in synaptic transmission and synaptic plasticity remains controversial. Perhaps the strongest case for a role of nitric oxide in neuronal signaling in the CNS is for long-term depression of synaptic transmission in the cerebellum.

Endocannabinoids

The primary psychoactive ingredient in cannabis, ⁹-tetrahydrocannabinol (⁹-THC), affects the brain mainly by activating a specific cannabinoid receptor, CB₁. CB₁ receptors are expressed at high levels in many brain regions, and they are primarily located on presynaptic terminals. Several endogenous brain lipids, including anandamide and 2-arachidonylglycerol (2-AG), have been identified as CB₁ ligands. These ligands are not stored, as are classic neurotransmitters, but instead are rapidly synthesized by neurons in response to depolarization and consequent calcium influx. Activation of metabotropic receptors (eg, by acetylcholine and glutamate) can also activate the formation of 2-AG. In further contradistinction to classic neurotransmitters, endogenous cannabinoids can function as retrograde synaptic messengers: They are released from postsynaptic neurons and travel backward across synapses, activating CB₁ receptors on presynaptic neurons and suppressing transmitter release. This suppression can be transient or long-lasting, depending on the pattern of activity. Cannabinoids may affect memory, cognition, and pain perception by this mechanism.

References