The following article from the NEJM might be of interest here
The New England Journal of Medicine -- April 19, 2001 -- Vol. 344, No. 16
Clinical implications of basic research
Anxiety at the Frontier of Molecular Medicine
Anxiety is a ubiquitous and unavoidable experience of life. It can be adaptive but also debilitating. Anxiety involves subjective
feelings (e.g., worry and a sense of threat), physiological responses (e.g., tachycardia and hypercortisolemia), and behavioral
responses (e.g., avoidance and withdrawal). Anxiety and fear share many subjective and physiological characteristics, and there
is much debate about how best to distinguish them. Fear, as characterized by Walter Cannon in the 1920s, is generally viewed as
a reaction to danger, whereas anxiety is a feeling of fear that is out of proportion to any real threat. When anxiety is persistent
and intrusive, it is referred to as generalized anxiety disorder. When it is manifested as sudden and repeated episodes of panic, it
is called panic disorder.
Anxiety is often a prominent manifestation of depression, trauma-related stress disorders, and certain personality disorders, such
as obsessive-compulsive disorder. According to Freudian psychology, anxiety is a signal of intrapsychic conflict, usually
between an unconscious wish and a learned prohibition. Research during the past decade has elucidated the neuronal circuitry
and molecular biology that underlie many of the manifestations of anxiety and the actions of anxiolytic medications.
Fear and behavior resembling anxiety are studied in animal models, which usually entail stress in the form of exposure to a
potentially dangerous environment (e.g., an elevated open platform), avoidance of noxious stimuli, and learned associations
between neutral stimuli and noxious stimuli. Stimuli that are perceived as potentially dangerous on the basis of prior learning
(e.g., conditioned fear) are processed by pathways leading from the thalamus and sensory cortex to specific limbic structures,
particularly the amygdala and hippocampus, which are critical for the initiation and mediation of learned responses to fear. (1)
Monkeys with lesions of the amygdala no longer display a fear of snakes; cats with such lesions show no fear of dogs. Humans
with damage to the amygdala have difficulty perceiving fear on someone's face and do not learn normally to identify stimuli that
signal danger. A recent functional neurologic imaging study has shown that physiologic activation of the amygdala occurs when
a subject is shown fearful faces even if the subject has no conscious awareness of having seen such a face. (2)
Specific neurons of the amygdala and hippocampus project widely to groups of cells that participate in the physiological and
behavioral responses associated with fear and anxiety. These cells occur in regions of the hypothalamus and brain stem that
mediate sympathetic and parasympathetic autonomic responses, the area of the paraventricular hypothalamic nucleus that
activates the glucocorticoid system, and groups of monoaminergic cells in the brain stem that are involved in arousal and priming
behavioral responses. The neurotransmitters of these pathways include glutamate, (gamma)-aminobutyric acid (GABA),
serotonin, norepinephrine, and the neuroactive peptide corticotropin-releasing factor.
Abnormal fear and behavior resembling anxiety are exhibited by mice that, through genetic engineering, lack one of the genes
that encode receptors implicated in mediating neurotransmission in these circuits, particularly serotonin 1A receptors,
corticotropin-releasing factor receptors, and specific subtypes of receptors for GABA. Other studies in animals have shown that
maternal behavior also affects fear circuitry. The more infant rats are actively nurtured by their mothers, the less extreme their
behavioral and molecular responses to fearful stimuli in adulthood. (3)
Of the various neurochemical agents used to treat symptoms of anxiety, benzodiazepines are the most broadly effective. The
brain has specific binding sites for benzodiazepines, and the actions of these agents are mediated at GABAA receptors, where
they potentiate the inhibitory effects of GABA. In animals, the infusion of benzodiazepines directly into the amygdala blocks fear
conditioning and the physiological response to perceived danger. However, because GABAA receptors are widely distributed in
the brain, benzodiazepines modify the effects of GABA in many brain circuits and therefore have diverse effects, such as
sedative or hypnotic effects, anticonvulsant actions, and muscle-relaxant effects; they can also cause ataxia and amnesia.
GABAA receptors are ligand-gated ion channels that mediate fast synaptic inhibition throughout the brain. The conformation of
the receptor changes when it binds to GABA: the channel opens, the inward chloride current increases, and the membrane
becomes hyperpolarized. The receptor is a pentameric, transmembrane glycoprotein composed of various combinations of 16
genetically distinct peptide subunits ((alpha), (beta), (gamma), (delta), (epsilon), (pi), and (rho) in all their isoforms). The specific
combination of subunits and isoforms varies widely throughout the brain, accounting for the diverse effects of GABA and of
drugs that affect GABAA receptors; however, most GABAA receptors consist of (alpha), (beta), and (gamma) subunits.
GABAA receptors mediate the central nervous system effects of benzodiazepines, barbiturates, alcohol, picrotoxin, inhaled
anesthetics such as isoflurane, penicillin, and certain neuroactive steroids. Most of these agents do not directly affect channel
conductance but rather allosterically modify the effect of GABA on the dynamics of channel opening. Benzodiazepines, for
instance, increase the frequency of channel openings induced by GABA. Combinations of (alpha) and (beta) subunits are
necessary to form an active GABA-binding site. To bind benzodiazepines these receptors require (alpha) and (gamma) subunits
-- specifically, several amino acid segments in the N-terminal extracellular domains of the (alpha) subunits. Moreover,
benzodiazepines will bind to these domains only if the (alpha) isoforms are types 1, 2, 3, or 5.
Studies involving tissue culture and genetically engineered mice have begun to identify the molecular specificity responsible for
the diverse effects of benzodiazepines. All four (alpha) subunits that bind to benzodiazepines have a conserved histidine residue
expressed in the N-terminal domain. GABAA receptors composed of (alpha)4 and (alpha)6 subunits, which do not bind or
respond to benzodiazepines, have an arginine at this residue (Figure 1). Inserting a point mutation that converts histidine to
arginine at this site causes receptors that are sensitive to become insensitive. Because the histidine residue seems to be critical
for the response to benzodiazepines, strains of mice have been created that have the His-to-Arg mutation in selective (alpha)
subunits in an effort to determine the molecular specificity of the effects of benzodiazepines. Mice with His-to-Arg mutations
only in (alpha)1 subunits, which are expressed abundantly throughout the cortex, show resistance to the sedative and amnesic
effects of classic benzodiazepines. Zolpidem, a hypnotic benzodiazepine with (alpha)1-specific binding, has no effects in the
animals. Benzodiazepines also have diminished anticonvulsant effects in these mice. The myorelaxant, ataxic, alcohol-enhancing,
and antianxiety effects of benzodiazepines are largely unchanged. (4,5)
In contrast, animals with selective His-to-Arg mutations in (alpha)2 subunits, which are expressed in the hippocampus,
amygdala, and cortex on the initial axonal segments of pyramidal neurons, show selective resistance to the antianxiety effects of
benzodiazepines but not to the sedative, motor, or anticonvulsant effects of these agents. (6) Mice with selective His-to-Arg
mutations in (alpha)3 subunits have none of the resistance evident in the other two strains.
These remarkable findings indicate that, at least in mice, (alpha)1 subunits are critical for mediating the sedative and probably
hypnotic effects of benzodiazepines, as well as their amnesic and, to a lesser degree, anticonvulsant effects. In contrast, (alpha)2
subunits appear preferentially to mediate the apparent antianxiety effect of these agents. The (alpha)3 and (alpha)5 subunits are
most likely involved primarily in the other actions of benzodiazepines. These observations suggest that new antianxiety agents
can be designed that will elicit only the desired effect and thus spare patients untoward effects. Moreover, the identification of
anxiety-specific pharmacology at the molecular level will lead to a more refined understanding of the neurobiology and genetics
of anxiety disorders.
Daniel R. Weinberger, M.D.
National Institute of Mental Health
Bethesda, MD 20892
References
1. LeDoux J. Fear and the brain: where have we been, and where are we going? Biol Psychiatry 1998;44:1229-38.
2. Whalen PJ, Rauch SL, Etcoff NL, McInerney SC, Lee MB, Jenike MA. Masked presentations of emotional facial expressions
modulate amygdala activity without explicit knowledge. J Neurosci 1998;18:411-8.
3. Francis D, Diorio J, Liu D, Meaney MJ. Nongenomic transmission across generations of maternal behavior and stress
responses in the rat. Science 1999;286:1155-8.
4. Rudolph U, Crestani F, Benke D, et al. Benzodiazepine actions mediated by specific (gamma)-aminobutyric acidA receptor
subtypes. Nature 1999;401:796-800.
5. McKernan RM, Rosahl TW, Reynolds DS, et al. Sedative but not anxiolytic properties of benzodiazepines are mediated by the
GABAA receptor (alpha)1 subtype. Nat Neurosci 2000;3:587-92.
6. Low K, Crestani F, Keist R, et al. Molecular and neuronal substrate for the selective attenuation of anxiety. Science
2000;290:131-4.
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