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Aspects of the Biochemical Pharmacology of Psychotropic Drugs

By Daniel X. Freedman, 1966
Only a decade ago, psychopharmacology faced the future equipped more with prescient hopes than with substantive findings[1]. Drugs were promised as tools to reveal the coding by which neuro­chemical as well as neurobehavioral sequences were regulated. Endogenous systems related to stress were to be unmasked and their relationship to behavior pathology revealed.

Therapeutic drugs were to be discovered which — if they did not specifically re­verse behavior disorder— would at the least set into motion com­pensatory and inhibitory processes. Drugs thereby would directly or indirectly permit a more successful operation of those contin­gencies normally regulating behavior and keeping it within accep­table bounds.

Today, it is the expanding range of specific information which is perhaps overwhelming. Where neurophysiologists have been able sufficiently to map the intricate sequences of peripheral and central signals which comprise the controls for basic drive be­haviors such as eating and drinking, a central chemical coding has been revealed [2, 3, 4].

For example, directly applied in select hypothalamic areas, norepinephrine can produce eating and its precursor (dopamine) can produce the effect after a delay suffi­cient for synthesis of the active amine; adrenergic blocking agents can reverse or block the effect while acetylcholine in the same area will produce drinking behavior. These effects are contingent upon highly localized concentrations and are not obtained with parent­eral or even intraventricular injection. Small changes in the mole­cular structure of indole or catechol amines can produce differences in psychotomimetic patterns and potency[5]. With a sleight of the medicinal chemist's hand chlorpromazine becomes the antidepressant imipramine.

Further, drugs have changed our very concept of the way in which familiar behaviors such as se­dation are put together. Substituents on the phenothiazine struc­ture produce an alert patient who may show drug-induced restless­ness while there is an inhibition of overly activated behavior. Both behaviorally and physiologically the "sedation" or "depression" produced by reserpine is different at different biochemically de­fined periods[5, 6] following the administration of the drug; such effects differ from the "sedation" production by phenothiazines[7].

Apparently, the functional anatomy of familiar behavior patterns is far more complex at both the neural and chemical levels than would be indicated by our inexplicit terminology and while phar­macologists may not always practice this preaching, it is from de­tailed pharmacological and clinical studies that such distinctions have been made. In brief, these various structure-activity rela­tionships have not only fulfilled earlier promises; they also indi­cate that we had not sufficiently appreciated the different compo­nents, the regulatory and compensatory systems underlying ap­parently similar behavioral states. Nor did we anticipate the varie­ty of chemically dependent linkages — the associations and disso­ciations — of which the nervous system is capable.

Limitations of Biochemical Pharmacology

The basic — perhaps preposterous — question posed by psycho­active drugs is how it is that the biochemical change can become behaviorally manifest; in a rigorous sense there is an appalling distance between biochemical mechanisms and the particular substrata for perception and behavior. The question inherently entails a detailed study of intrinsic control systems at a number of different levels from the enzymatic to the psychosocial. Our pro­gress has been of an empirical nature and the rapid proliferation of findings has occurred precisely because we are mapping out the terrain and specifying mechanisms and pathways which, in fact, occur in nature. Yet it is to be doubted that the most extensive encyclopedia of biochemical findings would be sufficient to ex­plain drug-behavior correlations.

If we make a distinction between drug action expressed in biochemical language and drug effect (which is determined by multiple factors) we can agree that a biochemical explanation alone must be of limited value. Every drug acts essentially to fa­cilitate, replace, alter or compete with the special and ubiquitous cellular mechanisms which normally regulate body chemistry. Hence no drug can be confined to effects on behavior. Similarly no behavioral pattern is wholly contingent on drug action, whe­ther we are studying isolated heart muscle or psychological atti­tude.

The surrounding conditions influence the pattern of effects. Since every drug has more than one action and more than one site of action, since prior state, dosage and route and schedule of drug administration are critical parameters of drug action and effect, the study of each drug requires a careful mapping of significant events. We might recall that simplistic though clear explanations of the effects of psychotropic drugs were available even before the evidence of the past ten years was accumulated and we can con­tinue to expect annual ad hoc explanations from those adroit in the byways of metapharmacology. The fact, however, is that we know far more about peripheral adrenergic nerve and associated responses than we do about the special problems of brain neuro­chemistry and neurobehavioral effects.

Unless we can couple biochemical sequences to precisely defined sequences of physio­logical and behavioral responses, unless we sufficiently control and define the effects in which we are interested and more closely link periods of biochemical change with behavioral change, our progress must be restricted.

Advances in Amine Biochemistry

Apart from these caveats, the striking advance of the decade is the fact that at long last we have learned something of the 'de­tailed biochemistry of those amines[8] which must underlie fight and flight behaviors. With respect to the catecholamines a number of exotic and endogenous amines have been identified (epinine, octopamine, synephrine). Biochemists possess a range of tools with which to manipulate the various sequences in amine synthe­sis and inactivation. These tools may be applied easily in vitro and occasionally, with success, in vivo.

Essentially, the task of the body is to convert amino acids from dietary sources into bioactive amines (Figure 1). Amines such as acetylcholine, serotonin, his­tamine, norepinephrine and its precursor dopamine are highly po­tent biological substances which, in minute quantities, induce physiologic responses in a number of extraneural tissues, as well as in certain neural systems.
Figure 1 The amine can be synthesized in one location in the periphery and delivered through the circulation to another peripheral tissue; the brain, however, must largely synthesize its own amines from the precursor amino acids and does so in a regionally specific pattern. If we focus on the catecholamines, there are two salient features to their inactivation (Figure 2).


Figure 2
First, the major excretory products are amines and acids (or al­cohols). Second, prior to excretion there are two pathways: the amine may be O-methylated or not; therefore the end products of amine metabolism consist either of unchanged amines and acids or of 0-methylated amines and acids (in which one of the ring-hy­droxyl groups is 0-methylated).

In a sense, the basic vocabulary is now available and the task of the future is to establish the syn­tax — the rules regulating the activity of these amines under nor­mal and abnormal conditions and their relationship to energy[9, 10] and fatty acid metabolism [11] and to endocrine as well as neu­ral function. We shall later review the significance of some drug effects on the various synthetic and inactivating pathways. Essen­tially, it is:
  1. The pattern of amine metabolites.
  2. Rates of change of tissue levels and the concentrations of amines at various cellu­lar and subcellular sites.
  3. Manipulation of the various enzy­matic steps which provide the biochemical clues with which the pharmacologist can establish relationships to function.

Explanatory Notions and Experimental Definitions

The most generally useful explanatory notion and one which has an ancient and respectable history in pharmacology has held that the behavioral changes following psychotropic drugs are due to excesses and deficiencies of endogenous substances at critical brain receptor sites — or to direct drug effects — or to both. Exces­ses and deficiencies would be achieved by changes in supply and demand, by synthesis, utilization and destruction; ultimately by binding and release of available substances at the receptor.

These useful notions, while applied to psychotropic drugs a decade ago, nevertheless had to be put into operational terms. The altered balance of amines at a receptor was envisaged but empiri­cal measures were feasible only if large alterations in whole brain levels occurred. Monoamine oxidase (MA) inhibitors produced large increases in amine levels due to decreased destruction of the amines while reserpine caused drops of more than 50 percent. The effect of reserpine was due to a release of amines, presumably from binding sites in storage compartments, and to an impair­ment of the binding of newly synthesized amine. This observation was of exceptional interest, since binding and release in the ab­sence of fluorimetrically measurable quantities of the drug were linked to a period of altered effects [13].

Shore, et al. [12, 13] hypothesized that the now free and unbound (but probably unmeasur­able) amine was capable of exerting a prolonged effect. Yet apart from the fact that this could be true for a number of amines fol­lowing reserpine it was clear that this drug-induced phenomena could not represent the only form of binding and release in nature and that changed levels of this magnitude cannot be the only de­finition of functional excesses and deficiencies. For example, small elevations in levels of serotonin following the serotonin precursor (5-HTP) produced sedation [14, 15] and small increases (on the or­der of 20%) in serotonin levels were observed following LSD (16).

The question also arises as to whether the "normal" variation in levels (changes of approximately 10%) is due to nature or to ex­perimental error. In any event this variation made the measure­ment of small changes, such as those produced by LSD, precarious, although theoretically all of these small changes could equally re­present actual binding and release phenomena.

The problem of technique, therefore, was to find measures more responsive to the concepts and to localize these changed lev­els in terms of the cellular compartments in which they might be occurring. For example, with the simple technique employed by Giarman and Schanberg [17, 18] — differential centrifugation — it was shown that the particulate matter of the brain cells contained approximately 70% of the serotonin and this fraction showed the greatest depletion following the full effects of reserpine.

Associa­tion of other amines with the granular material of cells both in the periphery[19, 20] and the brain[21] had been previously de­monstrated. It was found that a variety of psychotropic drugs—whatever their effect on whole brain levels—changed the distri­bution of amines in the particulate and supernatant cellular frac­tions which represent a number of different subcellular compo­nents[17, 23, 27].

The notion was that this changed distribution occurred because of the effects of drugs on the normal equilibria of the amines within the cells and that the relative amounts of amines in such fractions following drug treatment were conse­quential [23, 24]. In other words, localization was as important as total levels which could represent only the net effect of a number of processes. This work emphasized the basic idea that rates of change and the "traffic" of amines were relevant to observed ef­fects. What would be required were more refined measures to re­flect these equilibria and shifts in intracellular concentrations. Thus both biochemical measures of amine levels and turnover rates as well as structural considerations would be necessary to decode the "syntax" governing the operation of the biogenic amines.

Structure-integrated Function

There has, in fact, been a rapid development in techniques in the broad field of biochemical pharmacology as well as in the area of psychotropic drugs: radioisotopes, electron and fluorescence mi­croscopy, density gradient separation and differential centrifuga­tion — all of which are bringing biochemical sequences into closer approximation with the visualizable reality of structure.

The old concept that the cell is little more than a bag chock-full of enzymes has been replaced by a picture of a complex subcellular ultrastruc­ture. This is rapidly becoming biochemically mapped and evacuat­ed in terms of the implication for function of the structural locali­zation of chemical mechanisms. Coupled with our awareness of the regional localization in the brain of bioactive substances, this morphologic emphasis is the hallmark of contemporary work and represents the basis for current interpretations. The key for con­temporary biochemical pharmacology is, as Heinrich Waelsch once put it, "structure-integrated function."

Transport of Drugs to Brain Sites

Figure 3 is a diagram of a typical capillary-glial-neuronal assembly and significant subcellular elements (the specialized nerve ending with vesicles, the thickened presynaptic and post-synaptic patches of membrane, mitochondria scattered through­out the channels of endoplasmic reticulum). Unlike most tissues, the brain has little extracellular space and the perivascular glial cells interspersed between blood and neurone may serve as a spe­cialized exchange station analogous to an extracellular space.


Figure 3 Here, then, is a structural basis for the highly selective — often en­ergy-dependent—exchange of substances between blood and brain conceived of as the "blood-brain barrier." For instance, the selec­tive localization of carbonic anhydrase in glial cells may reflect a role in exchanging carbon dioxide between neuron and blood, and inhibitors of carbonic anhydrase (such as Diamox) may ameliorate certain seizure disorders in childhood. The ultrastructure of cap­illaries differs in the brain and this allows for differential transport in different regions[25].

The crucial question in tracing the impact of a drug on the critical bioactive sequences of the body—including the question of lipid solubility and selective transport —is the way in which drugs become accessible to active sites. In general, the amounts of drug required to exert an effect at an active receptor are far less than the amounts required to circumvent the liver and bring drug and receptor into contact. It appears that functionally significant brain regions preferentially concentrate chlorpromazine[26] (or LSD as recently reported by Snyder, et al.[27] and confirmed in our labo­ratories (28)), but extensive knowledge about the concentration and distribution of drugs in the brain is generally lacking.

Similarly (with some interesting exceptions) studies of drug metabolism as reflected in blood and urinary measures have not as yet provided a correlation with pharmacologic effects or with individual differ­ences in therapeutic response. It should be emphasized that blood levels of drugs as well as brain levels of drugs must be important since the chemoreceptors in such structures as the carotid sinus are potent in neurophysiologic regulations[29] and since certain specialized areas of the brain (such as the area postrema and the intercolumnar tubercles) are highly vascularized and may preferentially concentrate a drug. Such concentration as well as dif­ferential uptake and release of a drug at various brain areas must be accounted for in linking the sequence of drug, chemical and be­havioral changes.

The Psychotropic Drugs and Energy Metabolism

Drug effects on overall energy metabolism have not generally proved to be differentiating and the overall (as distinct from the regional) oxygen consumption of the brain appears mainly to de­fine the limits for coma or consciousness[30]. The mitochondrion consists of highly organized enzyme complexes integrated into an enclosed system of membranes — an arrangement permitting the efficient trapping of energy released during the oxidation of glu­cose. Phenothiazines can inhibit electron transport or uncouple this energy system from the oxidative processes (involving glucose intermediates) through which ADP is phosphorylated to yield/ATP. Yet the phenothiazine tranquilizers do not appear to work by reduc­ing energy supplies stored in ATP; ATP levels, at least in the whole brain, may in fact surprisingly increase after phenothiazine treat­ment [31].

Chlorpromazine does, however, quite sensitively inhi­bit the transition from resting oxidative states to states of high activity in brain slices and in isolated mitochondria [32, 33]. A more general effect of the phenothiazines is their ability to alter the configuration and permeability of various membranes and thus po­tentially to influence intercompartmental traffic (34). Speaking far too broadly, one can see that at many levels the effects of phen­othiazines are to stabilize an ongoing state; they dampen, for ex­ample, the often damaging and overcompensatory physiological responses occurring to certain stressors, by virtue of an action on interneural transmission in certain sites in the brain stem (35, 36).

Neuromodulators and Transmitters

The chief point of convergence in the attempt to link physico­chemical phenomena and bodily response has been the synapse where drugs must somehow influence the transfer of information between neurones. A specialized presynaptic structure — the nerve ending—contains "synaptic vesicles" which because of their proximity to the synaptic membrane are thought to be the storage sites allowing for efficient release — or retention of neurohumors; in the periphery, for example, the number of vesicles decreases after stimulation. The presumed transmitter would attach to syn­aptic membranes and initiate depolarization, or acting as a neuro­modulator, the substance would modify the environment in which the transmitter acts[37].

While norepinephrine is probably the per­ipheral adrenergic transmitter and while. there is some presump­tive evidence for acetylcholine as a transmitter in the brain, it is generally agreed[38] that we know too little, in fact, to engage in more than guessing games concerning the role of individual brain amines in synaptic transmission or in various neurobehavioral "functions." For purposes of orientation only, we can note that some, though not all[39], central effects of serotonin tend to be of a sedative nature and possibly some serotonin effects involve cho­linergic receptors; excesses of norepinephrine may be crudely associated with certain excitatory states and deficiencies with cer­tain states of exhaustion and sedation.

Levels of acetylcholine tend to correlate with physiologic sleep and excitation. Single-cell recording following electrophoretic administration of chemicals to brain reveal serotonin, norepinephrine and acetylcholine sensi­tive cells[42], and with microperfusion techniques, acetylcholine in the brain can be recovered in the effluent following stimulation[40, 41].

It should, however, be clear that simply because activity can be correlated with application or concentration of a substance in the brain, the substance cannot thereby be directly implicated as a synaptic transmitter or inhibitor. Nevertheless, the imaginative notion put forth by Fatt and Katz[43] that stored quanta of a trans­mitter would be released on nerve stimulation has been bolstered by evidence, including the fact that a number of endogenous a­mines have been found to be stored in vesicles (such as those vi­sualized in nerve endings); these substructures seem to have a capacity for concentrating exogenous amine [44, 45, 46, 47, 48]. With labeled amines, the uptake of radioactive grains associated with vesicles in sympathetic nerve has been actually visualized (49). With further refinements in technique such findings may yet be more precisely interpreted.

Intact peripheral nerve is required for the uptake and binding of amines and nerve section produces a loss of the vesicular bind­ing sites and supersensitivity of the receptor to injected amine (50). In the peripheral nerve, electrical stimulation (or certain drugs) will produce an efflux of amines [50, 51, 52].

Similarly, section and degeneration of various tracks in the central nervous system has been shown to produce changes in amine levels and in synthesiz­ing enzymes[53, 54, 55]. Elucidation of the relationship betWeen neural activity and brain enzymes and substrates, and the effect of central neural activity of hormones[56] and peptides on binding and release should provide the next major development in bio­chemical aspects of psychopharmacology.

The Life Cycle of Amines

Figure 4 provides a diagrammatic representation of the life cycle of the brain amines. Psychoactive drugs may act at a num­ber of points in this cycle: the transport of precursors, the synthesis in the cytoplasm of amines, their binding and release in subcellular structures — and their subsequent rebinding — or their enzymatic inactivation. Binding usually means functional inactivation or inaccessability of the amine due to storage in organelles; physi­cally, however, it can occur at the receptor or with soluble proteins in the cytoplasm and such "bound" amines could be functionally associated with activity or not.

Figure 1 How amines normally get bound or released is not known. The notion that there are pumps, not only at the cell wall for the brain amines but further pumps at the or­ganelles, has yet to be firmly proved. Much work remains to be ac­complished — especially in the brain; but when the chemist can lo­calize an enzyme in an organelle (e.g., dopamine beta oxidase in granules[57], he can then tell whether or not an amine has en­tered the granule or been shunted elsewhere, and when he can i­dentify hydroxylated metabolites he can deduce, theoretically, where the amine must have been to have undergone the bio­transformation.

Some 'sketches' of Amine Traffic

A current view of data is that there are at least two pools of bound amines. The pool conceived to be nearer the receptor repre­sents an easily released and more active pool. Sympathomimetic drugs or amines release this pool and thus indirectly cause hyper­tensive effects. After repeated dosage the pool is depleted and tachyphylaxis is observed. Catecholamines releasable in this way are thought to be inactivated by catechol-O-methyl transferase (COMT), an enzyme which Axelrod pictures as located in the cy­toplasm or extracellular space (58). Drugs such as reserpine in­duce release also from the deeper lying pool and these amines are shunted to the mitochondria' MAO and inactivated within the cell[59].

Following reserpine, inactive metabolites may cross the syn­aptic membrane and, accordingly, there is no hypertensive re­sponse. It may also be that reserpine affects the cell wall[60].

MAO inhibitors sometimes cause hypotension, even though actual levels of norepinephrine are increased. Apparently, these in­hibitors alone do not produce a functional excess of norepine­phrine. It is possible that the activity of COMT protects the recep­tor (and opinion varies about an additional effect of MAO inhibi­tors: inhibition of the release of amines [58, 61, 62, 68].

In any event, following MAO inhibition the easily-released pool may be still readily triggered by drugs or by amines (such as tyramine found in cheese) and this leads to quite potent hypertensive ef­fects. This may be due to the greater quantities of amine packed in the pools and 'ready' to be released by appropriate chemicals.

Chlorpromazine permits the storage of intracellularly syn­thesized amine but blocks the uptake of exogenous amines and the reuptake and recirculation of intracellular amines[58]. It there­by exposes amines more rapidly either to metabolism by COMT or to the receptors[63]; because chlorpromazine also blocks the re­ceptor, the overall effects are hypotensive.

Imipramine has simi­lar effects but the receptor is not blocked. The effects of imipra­mine and its active intermediate, desmethyl imipramine (DMI) are markedly enhanced when levels of catechols are changing —as during reserpine-induced release of norepinephrine (64). The mechanism by which imipramine "sensitizes" amine receptors is not clear[65].

Generally, both categories of antidepressant drugs do not seem to be directly acting, but rather to be sensitizing com­pounds and their activation effect in clinical depression may de­pend on the status of endogenous chemical releasers — the effects of which they markedly enhance. The view of antidepressants as sensitizing emphasizes the importance of prior neurobehavioral states upon drug effects. When depression is viewed behaviorally, it is as if a concurrent tension were converted into activity in the presence of these drugs; the functional status of the brain amines in depression becomes, therefore, an intriguing question.

Drugs such as alpha methyldopa (Aldomet) might be called "displacers"[66]. They and naturally-occurring amino acids compete for the synthesizing mechanism and are themselves converted to amines and stored in granules and released. Hence this leads to the notion of "false transmitters"[67, 68]. Other amino acids­phenylalanine compete with the serotonin precursor for uptake at the cell wall leading to altered brain amine levels, possibly of significance in phenylketonuria[69, 70].

Schematic representation of the typical capillary-glial-neuronal arrangement. A glial cell is seen interposed betweetrcapillary and neuron. An axon (AX) with its specialized nerve ending (NE), containing synaptic vesicles (SV) and mitochondria (MI), terminates at the neuron to form a synapse. An extensive system of internal mem­branes and channels, the endoplasmic reticulum (ER), is present in the neuron and, to a lesser extent, in the glial cells. Mitochon­dria are scattered throughout intervening areas of cytoplasm (stippled). Other abbreviations: CL: capillary lumen; EC: endo­thelial cell; GC: golgi complex; NU: nucleus; SC: synaptic cleft.

These sketches of the intricacies governing amine traffic (which were first indicated by differential centrifugation studies) are based largely on investigations from the laboratories of Julius Axelrod and those of Irving Kopin at the NIMH, where the fate of small amounts of highly labeled injected amine in peripheral tis­sues is investigated. However, the details of such schemes are fi­nally adjudicated; experimental studies show that a gamut of psy­chotropic drugs — antidepressants, phenothiazines and synpatho­mimetic amines—could influence the balance of amines at an ac­tive receptor.

Such studies have also shown that classical tools in biochemistry can be integrated with structural considerations. In summary, it is now generally accepted that psychotropic drugs in­fluence not only levels of amines, but their movements, sub-cellular localization, rates of turnover and accessibility, not only to receptors and to sites of storage, but to different pathways for enzymatic synthesis and destruction. Drugs may also block or in­duce enzymes. They may alter physical-chemical properties of membranes and receptor substance, influence ion flux, metabolic processes, or rates of binding and release and thus directly or in­directly influence neural functions.

Psychotomimetic Drugs

A number of active alkaloids isolated from plants and used ri­tually by natives (and even college students) to induce altered sub­jective states contain lysergic acid or structures related to biogenic indole and catechol amines (Figure 5). Figure 5 This group of compounds, drugs such as LSD-25, mescaline and psylocin and dimethyltry­ptamine (DMT) also influence binding and release of endogenous amines (5). Behaviorally, these substances produce a loss of the customary controlling anchors so that the usual boundaries which structure thought and perception become fluid; awareness be­comes vivid while control over input is markedly diminished. De­pendency either on the surroundings or on prior expectations or on a mystique for structure and support is enhanced.

Psychiatrists can recognize these primary changes as a background state out of which a number of secondary psychological states can ensue, depending on motive, capacity and circumstance. Our terminology reflects this: if symptoms ensue, the term psychotomimetic is em­ployed; and if mystical experience, religious conversion or thera­peutic behavior changes are stressed, the term psychedelic or "mind manifesting" has been applied. (And some thirty years ago chemists called the harmine- and harmaline-containing alkaloids, derived from South American cohaba, "telepathine").

The import­ance of these compounds does not lie essentially in an identity be­tween such drug-induced changes and the operative biochemical pathways in clinical disorders; rather, the drugs offer a tangible grip upon neurochemical sequences related to a peculiar and in­teresting mental state, and research with such a drug could pro­vide at least one detailed demonstration of how this state can, in fact, be achieved.

The question does arise as to whether mammals can produce endogenous psychotoxins and the enzymatic machin­ery for this appears to be present. For example, enzymes which can N-methylate indole or catechol amines are known; this means that DMT or bufotenine could be produced in the body. 6-hydroxy­lation of indoles can occur in the human and this substituent on the N-methyltryptamine structure enhances psychotoxicity in animals. The me thoxylation of catechols leads to a loss of sympath­omimetic potency but multiple methylation of phenethylamines leads to psychotoxic effects, as with mescaline. Friedhoff[71] has evidence that dimethyoxylation of catechols occurs more fre­quently, perhaps, in schizophrenics than normals. 5-methoxyla­tion of indoles heightens their behavioral effects and this enzyma­tic reaction occurs normally in the pineal, where N-acetyl seroto­nin is methoxylated to produce the skin-lightening hormone, me­latonin.

Beta carboline structures like harmaline are seen in an­other pineal hormone, adrenoglomeruloltropin, and Szara has pro­posed that such structures could be formed by the cyclizing of in-doles, perhaps as a psychoactive intermediate following drugs such as psilocin. Thus, demonstrations of the capacity of the human body to synthesize psychotoxic substances have clearly advanced, although it should be clear that such enzymatic conversions have not been etiologically linked with clinical disorders.

The relative potency of these compounds can be objectively demonstrated in rats working on a simple (FR) schedule in which the rat presses a lever for food[72]. With these measures LSD is about 150 times as potent as mescaline. The schedule can also de­monstrate a period of acute effects of LSD in the rat and the pheno­mena of tolerance and cross tolerance[73]. The acute effects fol­lowing an effective dose (ED90) of LSD in rats begins at five min­utes and are over by 30-45 minutes. As the LSD leaves the brain, and the serotonin is bound, these behavioral effects are observed (Figure 6).

Figure 6

We have recently found that the LSD at five minutes is concentrated in the particulate fraction and as it leaves this frac­tion and is concentrated in the supernatant, the binding of seroto­nin begins to occur in the particulate fraction, suggesting that LSD has changed the serotonin receptors there and defining a re­markably close relationship between LSD and the indole recep­tors [28].

There are further biochemical links between amine metabo­lism and LSD. Long after reserpine has been metabolized—or af­ter the effects of a short-acting releaser such as tetrabenazine are over (and amines are only 14% below normal) the effects of LSD in man and rat are enhanced and prolonged; doses of LSD in rats normally ineffective become potent [74, 75]. This rather surprising­ly proved not to be an effect of such drugs on the uptake of LSD in the brain since, in fact, less LSD enters the organ. Rather, there is enhanced duration of the binding of serotonin in the same cellu­lar fraction from which the reserpine and tetrabenazine release amines[28].

A number of psychotomimetic drugs bind serotonin[76] and release norepinephrine and such effects have been found with toxic dosage schedules of amphetamine[77]. Szara finds such effects most striking in the rabbit hypothalamus[78]. Preme­dication with certain MAO inhibitors may dampen the effects of LSD[79]. These and other[79, 80] studies, then, show that psycho­tomimetic drugs have an affinity for brain receptors which normal­ly process amines and that the binding of serotonin is somehow re­lated to a factor governing intensity of drug effect.

The presence of the drug in the body is also important. The half-life of LSD in plasma correlates with the duration of intense effects not only in man[81] but in rat, rabbit and cat, and we have some evidence that during the course of an LSD episode, fluctua­tions in plasma levels may correlate with periods of intensifica­tion of the LSD effect[28]. So that in spite of low drug levels (a few millimicrograms/cc in plasma) there may be several select re­ceptor sites in the brain which can continue to be stimulated by only a few molecules of drug held in the plasma during the periods of changing behavioral and amine effects.

Very low doses of LSD can produce behavioral effects in man[82] and animals[83] and biochemical correlates of these effects have yet to be found. Sco­polamine and piperydil glyocollates (Ditran) have no effects on the indole and catecholamines, but induce a 30% drop in levels of brain acetylcholine (84). Therefore, there are several groups of psychotomimetic drugs, both in terms of their behavioral and their chemical effects.

Synthesis and Destruction of Amines

Since the brain must synthesize amine from precursors, pre­cursors have been used not only to test the effects of excessive amine synthesized in situ, but to deduce the "real" effect of the amines. In view of the consequences of compartments for function, this is not an uncomplicated approach, especially if precursors are used after reserpine or MAO inhibitors.

Some drugs used in psy­chiatry influence various steps in the synthesis of amines (Figure 1); e. g. Antabuse inhibits dopamine beta oxidase. Recently it was reported that norepinephrine synthesis can be blocked at an early step (tyrosine hydroxylase) by alpha methyl tyrosine. This import­ant study by Spector, Sjoerdsma and Udenfriend[85] suggests that after inhibition we can give precursors later in the se­quence of norepinephrine synthesis (e. g., at 3 in Figure 1) and thus explore the role of dopamine in the central nervous system.

Further this inhibition permits the study of utilization — or physiological demand for the amine in various functional conditions — a key to many puzzles concerning the role of catecholamines. Finally the inhibition of synthesis by this method does not alter the binding, release and uptake mechanisms and hence various precursors and sympathomimetic drugs will not have bizarre effects, as observed following drugs which do alter amine regulations (such as MAO in­hibitors). The behavioral picture appears to be one of sedation, but a different pattern than that seen with reserpine. The animal ap­pears mainly to be easily fatigued and unable to withstand excita­tion or temperature extremes.

While the liver 0-methylates much of the circulating norepine­phrine (and hence the 0-methylated acid is a predominant urinary metabolite), the effect of drugs on various pathways of amine me­tabolism in Tissues can nevertheless be monitored (Figure 2). If drugs affect not only enzymes but accessibility of enzyme and sub­strate—the traffic of amines—we should and do see shifts in meta­bolic patterns following psychotropic drugs. Thus, following the release of amines by reserpine there is an increase in acid secre­tion which is followed by a decrease when storage is no longer possible[86].

Chlorpromazine, which prevents the uptake and re-uptake of the amines — perhaps exposing them to COMT but not MAO — produces a decrease in the 0-methylated acid (86). Follow­ing MAO inhibitors there is a marked decrease in the VMA excre­tion and it has recently been reported that there is a decrease fol­lowing imipramine (87). These reductions in acid by two antide­pressants occur, of course, with different mechanisms: one by en­zyme inhibition, the other by a shunting mechanism.

It should be remembered that accurate and fine measurement of even one me tabolite is a formidable methodological problem. What we now re­quire is a gamut of urinary and tissue metabolites to be screened after drugs. When this can be accomplished, it will be possible to deduce, more accurately, tissue events from urinary patterns. Metabolic patterns in the brain, of course, need not be reflected in urinary patterns; nevertheless, there are enzyme defects which are thought to be localized to the brain and in which the urine does produce concordant findings (88). We should soon have an accu­rate approximation of the contribution of brain metabolism to uri­nary metabolites. Some estimates are of the order of 20 percent.

Correlations of Metabolism with Stress

Patterns of urinary metabolites may reflect altered amine traf­fic as well as enzyme changes, but they have not clearly pointed to etiological factors. A welcome and significant change in emphasis has been to correlate changes in the pattern of endocrine or amine metabolites with side effects of drugs (89) or with functional changes occurring in the course of a behavior disorder. If the amines were indeed related to various intensity factors in beha­vior such a strategy would be more relevant to the behavioral facts than attempts to correlate classical nosology for clinical syndromes with metabolic change.

McDonald, et al.[86] found increased VMA excretion in chron­ic schizophrenic patients at the NIMH. This study implies an in­teresting definition of the stress which chronic schizophrenia may entail. When normal controls were exercising their usual activi­ties they showed increased norepinephrine metabolism, approach­ing, but not reaching, that of the patients. When the normals were in bed this increase disappeared, but the schizophrenic group still had an increased amine metabolism and in fact a slightly en­hanced metabolism.

Coupled with this observation is a study by Kornetsky, et al.[90] of this population. Postural hypotension was monitored twelve hours following a single dose of chlorpromazine. The patients showed minimal hypotension compared to normals. When the normals exercised for several hours, however, and a pos­tural hypotension test was then applied, the hypotension had for all practical purposes disappeared. In other words, the normal can be­gin to approximate the schizophrenic's response only with marked exertion; in spite of the apparent relative motor inactivity of the patients, there appears to be a great deal of activity of some kind in being a chronic schizophrenic.

The schizophrenics' active-inactivity or inactive-activity is reflected in an increased metabolism of norepinephrine which is, perhaps, greater even when the pa­tient is resting than when moving, and it is also evident in a rela­tive resistance to the marked hypotensive effects of chlorproma­zine. It has been clear for some time that if we are to study effects of stress in a manner relevant to clinical disorders, such as schizo­phrenia, we shall have to refine our notions of the physiological measures which may be appropriate and recognize and define the nature of the stress which is implied in psychological [91, 92] and psychotherapeutic studies of the disorder [93].

In general, the neu­ral and biochemical components of a variety of stressful states have yet to be disentangled and are poorly comprehended by con­fining attention solely to the adaptation syndrome. Since drugs act to influence binding and release we would have an elegant rationale for drug therapy, should there be a cor­responding pathophysiology of binding and release in the brain, either as a primary or secondary effect of psychosis or depression. There are no apparent direct approaches as yet to this question.

Although we know that chemical states affect neurobehavorial activity, in a basic sense our knowledge of how neurobehavoral activity affects chemical states is minimal [53,54,55]. It is, never­theless, of interest that with intense life threatening stress re­quiring marked exertion — such as the requirement that rats swim for their live — changes in brain amines occur and do so in the absence of the pituitary. These changes correlate with the ex­haustion and recovery of the animal — an instance of non-pharma­cologically induced change of brain amines[74].

From our own and other laboratories, there are now reports also of long-term changes seen both in brain catechol amines and acetylcholine in animals raised in various stressful environments[95,96]. Gillis has observed a shift in the subcellular distribution of norepine­phrine in the hearts of young rabbits contingent—simply on the injection of a needle into the peritoneum[97].

The pattern of brain amine response to acute exhaustion stress and to LSD in rats is strikingly similar. It is tempting to suggest that LSD — without inducing the physical exertion required by swimming — simply initiates a localized brain-stress response chemically. In other words, there are central components of ex­haustion stress which need not be coupled directly to physical activity. These central components could be activated by chem­icals or evoked by surtaxing exertion or extremes of temperature. In this analogy, schizophrenic patients would correspond to LSD activation and the normal subjects to the swimming rats. Local brainstress responses uncoupled from physical activity and reflected in changes of amine distribution or in vegetative responses could be compared to a motor racing with the clutch disengaged.

Finally, we should note a convergence of much more concrete evidence indicating that the brain amines normally may buffer the necessary psychophysiological regulations evoked by stimuli which induce an intense central activation. When we alter the controls normally governing binding and release (by pretreat­ment with MAO inhibitors or reserpine) and we then induce central activation with drugs or with amino acids (such as methio­nine or thyptophan) effects are produced which in humans are often described as psychotoxic[98] or in the case of MAO inhibi­tors and LSD, the expected psychotoxic effect may be lost. In other words, such pretreatment may have unmasked an underlying "buffer" role of the amines.

If genetic or stress induced impair­ment of binding and release occurred in clinical disorder, The human would be at a chemical disadvantage in handling life stresses and drugs would be helpful in compensating for these effects.

Leaving fantasy aside, we should note that the work of the future requires further detailed biochemical mapping of brain events and a linking of functional changes with biochemical measures. We still lack sufficient knowledge to have a basis for formal hypotheses which can be accepted or rejected by the critical experiment. For the behavioral and physiological scientist, the challenge is to describe more succinctly the sequences follow­ing drug administration, and to define and refine the functional units and behavioral dimensions appropriate for correlation with biochemical change.

Supported by United States Public Health Service grants MHK3-18, 566 and MH03363. Reprinted, by permission, from Freedman_, Daniel: Aspects of the Biochemical Itharrnacoloky Psychotropic Drugs. lw: Solomon, Philip (Ed.): Psychiatric Drugs. New York: Grune & Stratton,1966

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