GABA. Substantial data supports the idea that GABA mediates the inhibitory actions of local interneurons in the brain and may also mediate presynaptic inhibition within the spinal cord. Presumptive GABA-containing inhibitory synapses have been demonstrated most clearly between cerebellar Purkinje neurons and their targets in Deiter's nucleus; between small interneurons and the major output cells of the cerebellar cortex, olfactory bulb, cuneate nucleus, hippocampus, and lateral septal nucleus; and between the vestibular nucleus and trochlear motoneurons. GABA also mediates inhibition within the cerebral cortex and between the caudate nucleus and the substantia nigra. GABA-containing neurons and nerve terminals have been localized with immunocytochemical methods that visualize glutamic acid decarboxylase, the enzyme that catalyzes the synthesis of GABA from glutamic acid, or by in situ hybridization of the mRNA for this protein. GABA-containing neurons frequently co-express one or more neuropeptides.
The most useful compounds for confirmation of GABA-mediated effects have been bicuculline and picrotoxin (Figure 14–10); however, many convulsants whose actions previously were unexplained (including penicillin and pentylenetetrazol) are relatively selective antagonists of the action of GABA. Useful therapeutic effects have not yet been obtained through the use of agents that mimic GABA (such as muscimol), inhibit its active reuptake (such as 2,4-diaminobutyrate, nipecotic acid, and guvacine), or alter its turnover (such as aminooxyacetic acid).
GABA receptors have been divided into three main types: A, B, and C.
The most prominent GABA-receptor subtype, the GABAA receptor, is a ligand-gated Cl− ion channel, an "ionotropic receptor."
The GABAB receptor is a GPCR.
The GABAC receptor is a transmitter-gated Cl− channel.
GABAA receptor subunit proteins have been well characterized due to their abundance. The receptor also has been extensively characterized as the site of action of many neuroactive drugs, notably benzodiazepines, barbiturates, ethanol, anesthetic steroids, and volatile anesthetics (Figure 14–11).
Based on sequence homology to the first reported GABAA subunit cDNAs, multiple subunits have been cloned, including 6α, 4β, and 3γ subunits. They appear to be expressed in multiple multimeric, pharmacologically distinctive combinations. In addition to these subunits, which are products of separate genes, splice variants for several subunits have been described.
The GABAA receptor is probably pentameric or tetrameric in structure with subunits that assemble together around a central pore typical for other ionotropic receptors. The major form of the GABAA receptor contains at least three different subunits α, β, and γ, with likely stoichiometry of 2α, 2β, 1γ. All three subunits are required to interact with benzodiazepines with the profile expected of a native GABAA receptor. The distribution of different subunit combinations of GABAA and their functions in the mammalian brain are summarized in Table 14–2 (Fritschy and Mohler, 1995).
The GABAB or metabotropic GABA receptor interacts with Gi to inhibit adenylyl cyclase, activate K+ channels, and reduce Ca2+ conductance. Presynaptic GABAB receptors function as autoreceptors, inhibiting GABA release, and may play the same role on neurons releasing other transmitters. Functional GABA B receptors are heterodimers made up of GABABR1 and GABABR2 subunits (Bettler et al., 2004) The GABAC receptor is less widely distributed than the A and B subtypes. GABA is more potent by an order of magnitude at GABAC than at GABAA receptors, and a number of GABAA agonists (e.g., baclofen) and modulators (e.g., benzodiazepines and barbiturates) seem not to interact with GABAC receptors. GABAC receptors are found in the retina, spinal cord, superior colliculus, and pituitary (Olsen and Betz, 2005).
Glycine. Many of the features described for the GABAA receptor family also apply to the inhibitory glycine receptor, which is prominent in the brainstem and spinal cord. Multiple subunits assemble into a variety of glycine receptor subtypes. These pharmacological subtypes are detected in brain tissue with particular neuroanatomical and neurodevelopmental profiles. As with the GABAA receptor, the complete functional significance of glycine receptor subtypes is not known. There is evidence for clustering of glycine receptors by the anchoring protein gephyrin (Sola et al., 2004). An additional role for glycine is as a co-agonist at NMDA receptors, at which both glutamate and glycine must be present for activation to occur.
Glutamate and Aspartate. Glutamate and aspartate are found in very high concentrations in brain, and both amino acids have powerful excitatory effects on neurons in virtually every region of the CNS. Their widespread distribution initially obscured their roles as transmitters, but there now is broad acceptance that glutamate and possibly aspartate are the principal fast ("classical") excitatory transmitters throughout the CNS (Bleich et al., 2003; Conn, 2003). Multiple subtypes of receptors for excitatory amino acids have been cloned, expressed, and characterized pharmacologically, based on the relative potencies of synthetic agonists and the discovery of potent and selective antagonists (Kotecha and MacDonald, 2003). Glutamate receptors are classed functionally either as ligand-gated ion channel ("ionotropic") receptors or as "metabotropic" GPCRs (Table 14–3).
Neither the precise number of subunits that assembles to generate a functional glutamate ionotropic receptor ion channel in vivo nor the intramembranous topography of each subunit has been established unequivocally. The ligand-gated ion channels are further classified according to the identity of agonists that selectively activate each receptor subtype, and are broadly divided into N-methyl-D-aspartate (NMDA) receptors and non-NMDA receptors. The non-NMDA receptors include the α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), and kainic acid (KA) receptors (Table 14–3). Selective antagonists for these receptors are now available. In the case of NMDA receptors, agonists include open-channel blockers such as phencyclidine (PCP or "angel dust"), antagonists such as 5,7-dichlorokynurenic acid, which act at an allosteric glycine-binding site, and the novel antagonist ifenprodil, which may act as a closed-channel blocker. In addition, the activity of NMDA receptors is sensitive to pH and to modulation by a variety of endogenous agents including Zn2+, some neurosteroids, arachidonic acid, redox reagents, and polyamines such as spermine. Additional diversity of glutamate receptors arises by alternative splicing or by single-base editing of mRNAs encoding the receptors or receptor subunits. Alternative splicing has been described for metabotropic receptors and for subunits of NMDA, AMPA, and kainate receptors. For some subunits of AMPA and kainate receptors, the RNA sequence differs from the genomic sequence in a single codon of the receptor subunit that markedly affects the Ca2+ permeability of the receptor channel (Conn and Pin, 1997). AMPA and kainate receptors mediate fast depolarization at glutamatergic synapses in the brain and spinal cord. NMDA receptors are involved in normal synaptic transmission, but activation of NMDA receptors is usually associated more closely with the induction of various forms of synaptic plasticity rather than with fast point-to-point signaling in the brain. AMPA or kainate receptors and NMDA receptors may be co-localized at many glutamatergic synapses.
Activation of NMDA receptors is obligatory for the induction of a type of long-term potentiation (LTP) that occurs in the hippocampus. NMDA receptors normally are blocked by Mg2+ at resting membrane potentials. Thus, activation of NMDA receptors requires not only binding of synaptically released glutamate, but simultaneous depolarization of the postsynaptic membrane. This is achieved by activation of AMPA/kainate receptors at nearby synapses involving inputs from different neurons. AMPA receptors also are dynamically regulated to affect their sensitivity to the synergism with NMDA. Thus, NMDA receptors may function as coincidence detectors, being activated only when there is simultaneous firing of two or more neurons. A well-characterized phenomenon involving NMDA receptors is the induction of LTP. LTP refers to a prolonged (hours to days) increase in the size of a postsynaptic response to a presynaptic stimulus of given strength. NMDA receptors also can induce long-term depression (LTD; the converse of LTP) at CNS synapses. The frequency and pattern of synaptic stimulation may dictate whether a synapse undergoes LTP or LTD (Nestler et al., 2009). It is believed that NMDA-dependent LTP and LTD reflect the insertion and internalization of AMPA receptors, possibly mediated by CaM-kinase II and calcineurin/protein phosphatase 1, respectively (Figure 14–12).
Glutamate Excitotoxicity. High concentrations of glutamate lead to neuronal cell death by mechanisms that have only recently begun to be clarified (Figure 14–13). The cascade of events leading to neuronal death is thought to be triggered by excessive activation of NMDA or AMPA/kinase receptors, allowing significant influx of Ca2+ into neurons. Glutamate-mediated excitotoxicity may underlie the damage that occurs after ischemia or hypoglycemia in the brain, during which a massive release and impaired reuptake of glutamate in the synapse leads to excess stimulation of glutamate receptors and subsequent cell death. NMDA receptor antagonists can attenuate neuronal cell death induced by activation of these receptors (Haeberlein and Lipton, 2009). Because of their widespread distribution in the CNS, glutamate receptors have become targets for diverse therapeutic interventions. For example, a role for disordered glutamatergic transmission in the etiology of chronic neurodegenerative diseases and in schizophrenia has been postulated (Chapters 16 and 22).
Acetylcholine. Based on a non-homogeneous distribution within the CNS and the observation that peripheral cholinergic drugs could produce marked behavioral effects after central administration, many investigators addressed the possibility that ACh might also be a central neurotransmitter. In the late 1950s, Eccles and colleagues identified ACh as a neurotransmitter for the excitation of spinal cord Renshaw interneurons by the recurrent axon collaterals of spinal motoneurons. Subsequently, the capacity of ACh to elicit neuronal discharge has been replicated on scores of CNS cells (Shepherd, 2003). Eight major clusters of ACh neurons and their pathways have been characterized (Cooper et al., 2003; Nestler et al., 2009; Shepherd, 2003). For additional details, see Chapter 9.
In most regions of the CNS, the effects of ACh result from interaction with a mixture of nicotinic and muscarinic receptors. Nicotinic ACh receptors are found in autonomic ganglia, the adrenal gland, and in the CNS. Activation by ACh results in a rapid increase in the influx of Na+ and Ca2+ and subsequent depolarization. Nicotinic cholinergic receptors appear to desensitize rapidly. The receptors are composed by five heterologous subunits arranged around a central pore (see Figure 14–4). A total of 17 subunits, including 10 α, 4 β, as well as δ, ∊, and γ subunits, have been identified. The nicotinic receptor at the neuromuscular junction has the composition α2β∊δ. However, some α subunits, including α7, α8, and α9, can form functional homo-oligomers. For additional information on neuronal nicotinic ACh receptors, see Chapter 11.
There are five subtypes of muscarinic receptors, all of which are expressed in the brain. M1, M3, and M5 couple to Gq while the M2 and M4 receptors couple to Gi (Table 14–4). Several presumptive cholinergic pathways have been proposed in addition to that of the motoneuron-Renshaw cell.
Catecholamines. The brain contains separate neuronal systems that utilize three different catecholamines—dopamine, norepinephrine, and epinephrine. Each system is anatomically distinct and serves separate, functional roles within its field of innervation (Nestler et al., 2009)
Dopamine. Although DA was originally regarded only as a precursor of NE, assays of distinct regions of the CNS revealed that the distributions of DA and NE are markedly different. In fact, more than half the CNS content of catecholamine is DA and extremely large amounts of DA are found in the basal ganglia. There are three major DA-containing pathways in the CNS (Figure 14–14):
the nigrostriatal pathway
the mesocortical pathway, where neurons in the ventral tegmental nucleus project to a variety of midbrain structures and to the frontal cortex
the tuberoinfundibular pathway, which delivers DA to cells in the anterior pituitary
Initial pharmacological studies discriminated between two subtypes of DA receptors: D1 (which couples to GS to stimulate adenylyl cyclase) and D2 (which couples to Gi to inhibit adenylyl cyclase). Subsequent studies identified three additional genes encoding subtypes of DA receptors: the D5 receptor, which is related to the D1 receptor; and the D3 and D4 receptors, which are part of what is called the D2 receptor family. There are also two isoforms of the D2 receptor that differ in the predicted length of their third intracellular loops (Nestler et al., 2009). The D1 and D5 receptors activate the Gs-adenylyl cyclase-cyclic AMP-PKA system. The D2 receptors couple to multiple effector systems, including inhibition of adenylyl cyclase activity, suppression of Ca2+ currents, and activation of K+ currents. The effector systems to which the D3 and D4 receptors couple have not been unequivocally defined (Greengard, 2001) (Table 14–5). DA-containing pathways and receptors have been implicated in the pathophysiology of schizophrenia and Parkinson disease and in the side effects seen following pharmacotherapy of these disorders (Chapters 16 and 22).
Norepinephrine. There are relatively large amounts of NE within the hypothalamus and in certain parts of the limbic system, such as the central nucleus of the amygdala and the dentate gyrus of the hippocampus. However, this catecholamine also is present in significant, although lower, amounts in most brain regions. Detailed mapping studies indicate that noradrenergic neurons of the locus ceruleus innervate specific target cells in a large number of cortical, subcortical, and spinomedullary fields (Nestler et al., 2009). NE has been established as the transmitter at synapses between presumptive noradrenergic pathways and a wide variety of target neurons. For example, stimulation of the locus ceruleus depresses the spontaneous activity of target neurons in the cerebellum; this is associated with a slowly developing hyperpolarization and a decrease in membrane conductance (Aston-Jones et al., 2001; Nestler et al., 2009).
Multiple types and subtypes of adrenergic receptors (α1, α2, and β) and their subtypes have been described in the CNS; all are GPCRs (Table 14–6). As expected, β receptors couple to Gs and thence to adenylyl cyclase. α1 Adrenergic receptors are coupled to Gq, resulting in stimulation of phospholipase C, and are associated predominantly with neurons. α2 Adrenergic receptors are found on glial and vascular elements, as well as on neurons; they couple to Gi and thence to inhibition of adenylyl cyclase activity. α1 Receptors on noradrenergic target neurons of the neocortex and thalamus respond to NE with prazosin-sensitive, depolarizing responses due to decreases in K+ conductance. However, stimulation of α1 receptors also can augment cyclic AMP accumulation in neocortical slices in response to concurrent stimulation of the Gs pathway, possibly an example of Gq-GS cross-talk involving Ca2+/calmodulin and/or PKC (Ostrom et al., 2003). α2 Adrenergic receptors are prominent on noradrenergic neurons, where they presumably couple to Gi, inhibit adenylyl cyclase, and mediate a hyperpolarizing response due to enhancement of an inwardly rectifying K+ channel. As in the periphery, α2 receptors are located presynaptically, where they function as inhibitory autoreceptors. There is also evidence for postsynaptic α2 receptors that modulate sympathetic tone. Effects mediated through α2 receptors on blood pressure have been reported. It is believed, e.g., that the antihypertensive effects of the α2 selective agonist clonidine are due to stimulation of α2 receptors in the lower brainstem.
Epinephrine. Neurons in the CNS that contain epinephrine were recognized only after the development of sensitive enzymatic assays and immunocytochemical staining techniques for phenylethanolamine-N-methyltransferase, the enzyme that converts NE into epinephrine. Epinephrine-containing neurons are found in the medullary reticular formation and make restricted connections to pontine and diencephalic nuclei, eventually coursing as far rostrally as the paraventricular nucleus of the thalamus. Their physiological properties have not been unambiguously identified.
5-Hydroxytryptamine (Serotonin). In mammals, 5-HT containing neurons are found in nine nuclei lying in or adjacent to the midline (raphe) regions of the pons and upper brainstem. Cells receiving cytochemically demonstrable 5-HT input, such as the suprachiasmatic nucleus, ventrolateral geniculate body, amygdala, and hippocampus, exhibit a uniform and dense investment of serotinergic terminals.
Molecular biological approaches have led to identification of 14 distinct mammalian 5-HT receptor subtypes (Table 14–7). These subtypes exhibit characteristic ligand-binding profiles, couple to different intracellular signaling systems, exhibit subtype-specific distributions within the CNS, and mediate distinct behavioral effects of 5-HT. Most 5-HT receptors are GPCRs coupling to a variety of G-protein α subunits. The 5-HT3 receptor, however, is a ligand-gated ion channel with structural similarity to the α-subunit of the nicotinic acetylcholine receptor. As seen with subtypes of glutamate receptors, mRNA editing has also been observed for the 5-HT2C receptor (Niswender et al., 2001); the resulting isoforms differ in agonist affinity and distribution in the brain.
The family of 5-HT1 receptors is composed of at least five receptor subtypes (Table 14–7) that are linked to inhibition of adenylyl cyclase activity or to regulation of K+ or Ca2+ channels. 5-HT1A Receptors are abundantly expressed on 5-HT neurons in the dorsal raphe nucleus, where they are thought to be involved in temperature regulation. They also are found in regions of the CNS associated with mood and anxiety such as the hippocampus and amygdala. Activation of 5-HT1A receptors opens an inwardly rectifying K+ conductance, which leads to hyperpolarization and neuronal inhibition. These receptors can be activated by drugs including buspirone, which is used for the treatment of anxiety and also used off-label for panic disorders. In contrast, 5-HT1D receptors are activated by low concentrations ofsumatriptan, which is currently prescribed for acute management of migraine headaches (Chapters 13 and 34). The 5-HT2 receptor class has three subtypes: 5-HT2A, 5-HT2B, and 5-HT2C; these receptors couple to pertussis toxin-insensitive G proteins (e.g., Gq and G11) and link to activation of PLC. Based on ligand binding and mRNA in situ hybridization patterns, 5-HT2A receptors are enriched in forebrain regions such as the neocortex and olfactory tubercle, as well as in several nuclei arising from the brainstem. The 5-HT2C receptor, which is very similar in sequence and pharmacology to the 5-HT2A receptor, is expressed abundantly in the choroid plexus, where it may modulate cerebrospinal fluid production. Many of the effects of antidepressants are thought to be a consequence of increased stimulation of 5-HT receptors following inhibition of 5-HT reuptake by SERT. On the other hand, inhibition of 5-HT2c receptors may account for the increased weight gain associated with neuroleptics used to treat schizophrenia.
5-HT3 receptors function as ligand-gated ion channels; these receptors were first recognized in the peripheral autonomic nervous system. Within the CNS, they are expressed in the area postrema and solitary tract nucleus, where they couple to potent depolarizing responses that show rapid desensitization to continued 5-HT exposure. Actions of 5-HT at central 5-HT3 receptors can lead to emesis and anti-nociceptive actions, and 5-HT3 antagonists are beneficial in the management of chemotherapy-induced emesis (Chapter 46).
Within the CNS, 5-HT4 receptors occur on neurons within the inferior and superior colliculi and in the hippocampus. Activation of 5-HT4 receptors stimulates the Gs-adenylyl cyclase-cyclic AMP pathway. Other 5-HT receptors are less well studied in the CNS. The 5-HT6 and 5-HT7 receptors also couple to Gs; their affinity for clozapine may contribute to its antipsychotic efficacy (Chapter 16).
The hallucinogen lysergic acid diethylamide (LSD) is a potent partial agonist at 5-HT2 receptors. When applied iontophoretically, LSD inhibits the firing of raphe (5-HT) neurons. The inhibitory effect of LSD on raphe neurons offers a plausible explanation for its hallucinogenic effects, namely that these effects result from depression of activity in a system that tonically inhibits visual and other sensory inputs. However, typical LSD-induced behaviors are seen in animals with destroyed raphe nuclei or after blockade of the synthesis of 5-HT by p-chlorophenylalanine (Aghajanian and Marek, 1999); Nichols, 2004).
Histamine. Histamine and antihistamines have long been known to produce significant effects on animal behavior. Biochemical detection of histamine synthesis by neurons and direct cytochemical localization of these neurons have defined a histaminergic system in the CNS. Most of these neurons are located in the ventral posterior hypothalamus; they give rise to long ascending and descending tracts that are typical of the patterns characteristic of other aminergic systems. Based on the presumptive central effects of histamine antagonists, the histaminergic system is thought to affect arousal, body temperature, and vascular dynamics. Four subtypes of histamine receptors have been described; all are GPCRs (Figure 14–15). H1 receptors, the most prominent, are located on glia and vessels as well as on neurons and act to mobilize Ca2+ in receptive cells through the Gq-PLC pathway.
H2 receptors couple via GS to the activation of adenylyl cyclase. H3 receptors, which have the greatest sensitivity to histamine, are localized primarily in basal ganglia and olfactory regions in rat brain and act through Gi to inhibit adenylyl cyclase. Consequences of H3 receptor activation remain unresolved but may include reduced Ca2+ influx and feedback inhibition of transmitter synthesis and release (Chapter 32). H4 receptors are expressed on cells of hematopoietic origin: eosinophils, T cells, mast cells, basophils, and dendritic cells. H4 receptors appear to couple to Gi and Gq and are postulated to play a role in inflammation and chemotaxis (Thurmond et al., 2004). Unlike the monoamines and amino acid transmitters, there does not appear to be an active process for reuptake of histamine after its release. Inhibition of H1 receptors causes drowsiness, an effect that limits the use of H1 antagonists to treat allergic reactions. The development of H1 antagonists with low CNS penetration has reduced the incidence of these side effects.
Peptides. The discovery during the 1980s of numerous novel peptides in the CNS, each capable of regulating neuronal function, produced considerable excitement and an imposing catalog of substances as well as potential medications based upon interaction with peptide receptors (Darlison and Richter, 1999; Hökfelt et al., 2003). In addition, certain peptides previously thought to be restricted to the GI tract or to endocrine glands have been found in the CNS. Most of these peptides bind to GPCRs. Many of the effects are modulatory rather than causing direct excitation or inhibition. Myriad peptide neurotransmitters or neuromodulators have been described (Table 14–8). Relatively detailed neuronal maps are available that identify immunoreactivity to peptide-specific antisera. While some CNS peptides may function on their own, most are now thought to act primarily in concert with co-existing transmitters, both amines and amino acids.
Neuropeptides are processed and stored in large, dense-core vesicles (LDCVs; see Figure 14–7). The peptides may be co-localized and released together with small molecule transmitters, such as a biogenic amine. Multiple peptides may be co-localized within the same neuron. As noted, some neurons may contain two or more transmitters, including peptides, and their release can be independently regulated.
In contrast to the biogenic amines or amino acids, peptide synthesis requires transcription of DNA into mRNA and translation of mRNA into protein. This takes place primarily in perikaria and the resulting peptide is then transported to nerve terminals. Single genes can, through the post-translational action of peptidases, give rise to multiple neuropeptides. For example, proteolytic processing of proopiomelanocortin (POMC) gives rise to, among other things, ACTH, α and γ MSH, β-MSH, and β-endorphin (Figure 14–16). In addition, alternative splicing of RNA transcripts may result in distinct mRNA species. For example, calcitonin and calcitonin gene-related peptide (CGRP) are derived in specific tissues from the same primary transcript.
Organization by Function. Since most peptides were identified initially on the basis of bioassays, their names reflect these biologically assayed functions (e.g., thyrotropin-releasing hormone and vasoactive intestinal polypeptide). These names become trivial when more ubiquitous distributions and additional functions are discovered. Some general integrative role might be hypothesized for widely separated neurons (and other cells) that make the same peptide. However, a more parsimonious view is that each peptide has unique messenger roles at the cellular level that are used repeatedly in biologically similar pathways within functionally distinct systems.
Most neuropeptide receptors are GPCRs. In comparison to GPCRs for smaller ligands such as biogenic amines and amino acids, the extracellular domains of neuropeptide receptors play a larger role in ligand binding. As seen with other transmitter systems, there are often multiple subtypes of receptor for the same peptide transmitter (Table 14–9). For example, there are five subtypes of receptor for somatostatin and all inhibit adenylyl cyclase through an interaction with Gi; they differ in their interaction with various somatostatin analogs. The cloning of the major members of the opioid-peptide receptors has revealed unexpected, and as yet unexplained, homologies with receptors for somatostatin, angiotensin, and other peptides. Multiple melanocortin receptors exist that respond to various peptides derived from POMC. Not surprisingly, the receptor on adrenal cortical cells responds to ACTH, while that on melanocytes responds to α-MSH. Further evidence for complexity comes from the realization that agonists at subtypes of melanocortin receptors are associated with a variety of biological effects including skin darkening (MCR1), decreased appetite (MCR3 and/or MCR4), and sexual arousal (MCR4).
Although most peptide receptors are GPCRs, exceptions do exist. The amiloride-sensitive FMRF amide (phe-met-arg-phe-amide) receptor is a peptide-gated ion channel. The large number of neuropeptides and peptide receptors has lead to increased interest in identifying therapeutic agents. Examples include experimental substance P antagonists as antidepressants or to treat anxiety, and antagonists of CRH for stress related disorders.
Comparison with Other Transmitters. Peptides differ in several important respects from the monoamine and amino acid transmitters. Peptide synthesis takes place in the rough endoplasmic reticulum. The propeptide is cleaved (processed) to the secreted form as secretory vesicles are transported from the perinuclear cytoplasm to the nerve terminal. Active mechanisms for the local synthesis of peptides have not been described; thus, peptidergic nerve terminals depend on distant sites of synthesis. As for structure-activity relationships for peptide transmitters, linear chains of amino acids can assume many conformations at their receptors, making it difficult to define the sequences and their steric relationships that are critical for activity. Until recently, it was difficult to develop non-peptidic synthetic agonists or antagonists that interact with specific peptide receptors. Such agents now are being developed for many neuropeptides (Hökfelt et al., 2003). Natural products have not been good sources of drugs that affect peptidergic transmission. Only one plant alkaloid, morphine, has been found to act selectively at peptidergic synapses. Fortunately for pharmacologists, morphine was discovered before the endorphins, or rigid molecules capable of acting at peptide receptors might have been deemed impossible to develop.
Other Regulatory Substances.
Cannnabinoids. Delta-9-tetrahydrocannabinol (THC) is one of several active substances in marijuana (Figure 14–17). It has dramatic short-term effects, including causing feelings of euphoria and altered sensory perception. After chronic or long-term use, withdrawal symptoms include irritability and sleep disturbances. The primary pharmacologic effects of THC follow its interaction with CB1 receptors in the CNS and CB2 receptors in the periphery. CB1 receptors are found primarily in the basal ganglia, hippocampus, cerebellum, and cerebral cortex. They are also expressed in some non-neuronal cells and tissues, including leukocytes and testis. CB2 receptors are expressed in the spleen, tonsils, bone marrow, and on peripheral blood leukocytes. The natural endogenous ligands for these receptors are arachidonic acid derivatives including anandamide and 2-arachidonyl glycerol (Figure 14–17). Both CB1 and CB2 receptors are linked to Gi and inhibition of adenylyl cyclase activity. Activation of CB1 receptors results in inhibition of glutamate release. Efforts to develop CB1 antagonists like rimonabant have focused on possible treatments for drug addiction and obesity. Efforts are also underway to develop agonists that interact with CB1 and CB2 receptors for the relief of pain. THC (dronabinol, MARINOL) is sometimes used in the control of nausea and moderate pain (Chapter 46).
Purines. Adenosine, ATP, UDP, and UTP have roles as extracellular signaling molecules (Robertson et al., 2001; Siegel et al., 2006). High concentrations of ATP are found in adrenergic storage vesicles and ATP is released along with catecholamines and the other contents of adrenergic storage granules. Intracellular nucleotides may also reach the cell surface by other means (Lazarowski et al., 2003), including as a consequence of cellular hypoxia or cell death, and extracellular adenosine can result from cellular release and metabolism of ATP (Jackson and Raghvendra, 2004). The concentration of ATP may greatly exceed the concentration of adenosine and, given the presence of nucleotidases, it can be difficult to unambiguously distinguish between effects of, e.g., adenosine and ATP.
Extracellular nucleotides and adenosine act on a family of purinergic receptors that is divided into two classes, P1 and P2 (Table 14–10). P1 receptors are GPCRs that interact with adenosine; two of these receptors (A1 and A3) couple to Gi and two (A2a and A2b) couple to Gs; methylxanthines antagonize A1 and A3 receptors. Activation of A1 receptors is associated with inhibition of adenylyl cyclase, activation of K+ currents, and in some instances, with activation of PLC; stimulation of A2 receptors activates adenylyl cyclase. The P2 class includes a large number of P2X receptors that are ligand-gated ion channels and the P2Y receptors, a similarly large subclass of GPCRs that couple to Gq or Gi and their associated effectors. The P2Y14 receptor is expressed in the CNS; it interacts with UDP-glucose and may couple to Gq. The P2Y12 receptor is important clinically, since inhibition of this receptor in platelets inhibits platelet aggregation.
Although many of these receptors have been detected in brain, most of the current interest stems from pharmacological rather than physiological observations. Adenosine can act presynaptically throughout the cortex and hippocampal formation to inhibit the release of amine and amino acid transmitters. ATP-regulated responses have been linked pharmacologically to a variety of pathophysiological functions, including anxiety, stroke, and epilepsy. A1 antagonists are being investigated as potential therapeutic agents to enhance awareness and learning. A2 receptors and dopamine D2 receptors appear to be functionally antagonistic, leading to investigation of A2a antagonists as adjunctive therapy for Parkinson disease (Jacobson and Gao, 2006).
Lipid Mediators. Substances identified as physiological regulators in systems throughout the body have been examined for their roles within the CNS. Arachidonic acid, normally stored within the cell membrane as a glycerol ester, can be liberated during phospholipid hydrolysis (by pathways involving phospholipases A2, C, and D). Phospholipases are activated via a variety of receptors (Chapter 3). Arachidonic acid can be converted to highly reactive regulators by three major enzymatic pathways (Chapter 33): cyclooxygenases (leading to prostaglandins and thromboxane), lipoxygenases (leading to the leukotrienes and other transient catabolites of eicosatetraenoic acid), and CYPs (which are inducible and also expressed at low levels in brain). Arachidonic acid metabolites have been implicated as diffusible modulators in the CNS, possibly involved with the formation of LTP and other forms of neuronal plasticity.
Nitric Oxide and Carbon Monoxide. Nitric Oxide (NO), an important regulator of vascular and inflammatory mediation, came into focus with respect to roles in the CNS after the characterization of brain nitric oxide synthase (NOS) activities (see Chapter 3 and Boehning and Snyder, 2003). Both constitutive and inducible forms of NOS are expressed in the brain. The application of inhibitors of NOS (e.g., methylarginine and nitroarginine) and of NO donors (such as nitroprusside) suggests the involvement of NO in a host of CNS phenomena, including neurotransmitter release and enhancement of glutamate (NMDA)-mediated neurotoxicity and LTP. Neuronal NOS (nNOS) is activated following NMDA-mediated receptor mediated increases in intracellular Ca2+. NO diffuses freely across membranes, acting primarily to stimulate soluble guanylyl cyclase, which catalyzes the formation of cyclic GMP. Formation of NO can also result in S-nitrosylation of cysteine residues in a variety of proteins including G proteins and ion channels. Other gases may also act as intracellular messengers. One candidate is carbon monoxide (CO), which is generated in neurons by three isoforms of hemeoxygenase (HO), isoform 2 predominating in neurons. Like NO, CO stimulates soluble guanylyl cyclase.
Cytokines. Cytokines are a large and diverse family of polypeptide regulators that are produced widely throughout the body by cells of diverse embryological origin. The effects of cytokines are regulated by the conditions imposed by other cytokines, interacting as a network with variable effects leading to synergistic, additive, or opposing actions. Tissue-produced peptides, termed chemokines, serve to attract immune and inflammatory cells into interstitial spaces. These special cytokines have received attention as potential regulators of nervous system inflammation (as in early stages of dementia, following infection with human immunodeficiency virus, and during recovery from traumatic injury). Some of the more conventional neuronal- and glial-derived growth-enhancing and growth-retarding factors have been identified earlier. The fact that neurons and astrocytes may be induced under some pathophysiological conditions to express cytokines or other growth factors further blurs the dividing line between neurons and glia (Campbell, 2004; Wang et al., 2002).