Is cholinergic sensitivity a genetic marker for the affective disorders?

zy zyxw zyxwvuts zy American Journal of Medical Genetics (Neuropsychiatric Genetics) 54:33&344 (1994) Is Cholinergic Sensitivity a Genetic Marker for the Affective Disorders? David S. Janowsky, David H. Overstreet, and John I. Nurnberger, Jr. Center for Alcohol Studies and Department of Psychiatry, University of North Carolina School of Medicine, Chapel Hill, North Carolina (D.S.J., D.H.O.), and Institute for Psychiatric Research, Indiana University School of Medicine, Indianapolis, Indiana (J.I.N.) The recent literature on the involvement of cholinergic muscarinic mechanisms and adrenergidcholinergic balance in affective disorders is reviewed and integrated with the older literature. There is strong evidence supporting the presence of exaggerated responses (behavioral, neuroendocrine, sleep) to cholinergic agents in affective disorder patients relative to normal controls and certain other psychiatric patients. There is also some, albeit less, conclusive evidence that these exaggerated responses may occur in euthymic individuals with a history of affective disorders, or in children at risk for development of affective disorders. Despite these promising results, suggesting a role for acetylcholine in the genetics of the affective disorders, further work in biochemistry and genetics is needed to link specific muscarinic receptors or other cholinergic variables to affective illness. 0 1994 Wiley-Liss, Inc. link several disorders, while not necessarily producing similar symptom complexes. A number of biochemical alterations, including changes in central norepinephrine, dopamine, serotonin, beta endorphin, and GABA have been postulated as etiologic factors in the affective disorders. Alterations in several of these neurotransmitters appear to lead to symptom alleviation. Nevertheless, there has been a paucity of evidence showing that alterations occur in these neurotransmitters before and after manifest affective illness is present. Acetylcholine appears to be one neurochemical which indeed induces altered effects in affective disorder patients, whether the patients are symptomatic or asymptomatic. Increasing central acetylcholine causes depressive symptoms, alleviates manic symptoms, and a t the same time is consistently associated with exaggerated muscarinic responses in affective disorder patients [Janowsky and Risch, 19871. Thus, muscarinic cholinergic hyperactivity or hypersensitivity may be a marker of the affective disorders, and indeed, such hypersensitivity as such may be a marker of a genetic propensity to develop an affective disorder. The present review paper will critically examine evidence both in support of and contrary to the possibility that muscarinic cholinergic mechanisms and effects may represent potential genetic markers of the affective disorders. It will draw heavily from evidence accumulated over the past two decades exploring an adrenergic-cholinergic balance hypothesis of depression and mania, in which depression is considered due to a predominance of or a supersensitivity to acetylcholine, relative to aminergic mechanisms, and mania is considered the converse [Janowsky et al., 1972; Siever et al., 19811. zyxwvutsr KEY WORDS: acetylcholine, genetics, affective disorders, depression INTRODUCTION Although not unheard of, there have been few consistently reported biologic state markers of affective illness reported t o date, and even fewer trait markers. This fact has made difficult the search for genetic markers for the affective disorders, producing a need for reliance on applied phenomenologic diagnostic criteria. Furthermore, although diagnostic criteria may be relatively reliable, they do not take into consideration the possibility that underlying genetic factors may INCREASED PRESYNAPTIC CHOLINERGIC ACTMTY IN AFFECTIVE DISORDER PATIENTS The vast majority of evidence suggesting that increasing central acetylcholine can induce depression has come from the utilization of cholinergic agonist and cholinesterase inhibiting strategies (see following sections). Although strategies involving stimulation of central muscarinic mechanisms (i.e., using such cholinergic agents as arecoline, oxotremorine, DFP, and zyxwvutsrq Received for publication November 16, 1993; revision received March 7,1994. Address reprint requests to David S. Janowsky, M.D., Department of Psychiatry, CB #7160, University of North Carolina School of Medicine, Chapel Hill, NC 27599-7160. 0 1994 Wiley-Liss, Inc. 336 Janowsky et al. zyxwvuts physostigmine) have provided much evidence supporting a cholinergic alteration in the affective disorders, these observations have all been based on indirect evidence. These studies have provided no direct evidence with respect to presynaptic cholinergic function in the affective disorders. Clinical in vivo hydrogen magnetic resonance spectroscopy provides a means for more directly assessing human presynaptic cholinergic function in vivo. This technique can measure choline-containing substances noninvasively in the brain. Such measurement is significant because choline is the major precursor of acetylcholine throughout the nervous system. Using in vivo hydrogen magnetic resonance spectroscopy, Charles et al. 119931observed an increase in central choline, the rate-limiting precursor t o acetylcholine in the imaged brains of patients with major depression, as compared to controls. This increase in imaged choline reverted to normal levels after successful drug treatment of the patients’ depression. The study by Charles et al. therefore suggests that presynaptic cholinergic overactivity may be associated with depression. The fact that the choline levels in this study returned to normal in the recovered patients suggests that increased central choline may be a state marker, rather than a trait marker of depression. Much more work with this promising technique is needed before a definitive conclusion will be possible, but the data presently available are consistent with the hypothesis that increased central acetylcholine may be associated with the affective disorders. BEHAVIORAL PHENOMENA This section will review the several lines of evidence suggesting that symptoms exhibited naturalistically by depressive patients and those shown by normal and depressed individuals receiving cholinomimetic agents are similar, and that the behavioral symptoms exhibited by individuals in general after the administration of cholinergic agonists are exaggerated in depressed patients. Phenomenological Similarities Between Depression and the Effects of Cholinomimetics Probably the most convincing evidence that acetylcholine is involved in the regulation of the affective disorders is the observation that centrally active cholinomimetic drugs, which increase central acetylcholine levels or stimulate central muscarinic receptors, rapidly induce depressed moods (i.e., sadness, guilt feelings, crying, regretfulness, etc.), and antagonize manic symptoms [Janowsky et al., 1972, 1973a,b; Carroll et al., 1973; Shopsin et al., 1975; Davis et al., 19781. In addition to observations of depression-induction caused by DFP [Rowntree et al., 19501and insecticides [Gershon and Shaw, 19611, Janowsky et al. [19811 found induction andlor intensification of depressed mood in approximately 75% of those in groups of unipolar-depressed, schizoaffective-depressed, and manic patients, and in approximately 25% of normals. Similarly, Davis et al. [1978] and Modestin et al. [1973a,bl demonstrated antimanic effects and an increase in depression in manic patients given the cholinesterase in- hibitor, physostigmine. Risch et al. [1982,19831, studying depressed patients, and Nurnberger et al. [1983al, studying euthymic bipolar patients given the direct muscarinic cholinergic receptor agonist, arecoline, also developed depressed mood and other forms of negative affect. These included hostility and anxiety. Physostigmine was also reported to induce a depressed mood in a majority of euthymic bipolar patients maintained on lithium [Oppenheimer et al., 19791. Depression-like symptoms have also been observed in normals receiving intravenous physostigmine or arecoline [Risch et al., 1981a;Nurnberger et al., 1983a), in Alzheimer’s patients receiving the cholinergic agonist oxotremorine [Mohs et al., 19871, and in normals who had smoked marijuana after receiving physostigmine [El-Yousef et al., 1973; Davis et al., 19781. Thus, a wide range of individuals have exhibited depressed mood andlor other negative affects after administration of cholinergic agonists and cholinesterase inhibitors. Depressed moods have also been observed in subjects receiving acetylcholine precursors including deanol, choline, and lecithin. Davis et al. 119791 and Tamminga et al. [1976] found that depressive symptoms occurred in some schizophrenic patients who were treated with choline. In a subgroup of cases, it was noted that depressed mood was a side effect of choline and lecithin treatments employed to try to reverse the memory deficits of Alzheimer’s disease [Bajada, 19821. Also, Casey [1979] observed that a depressed mood and, in some cases, a paradoxical hypomania occurred in a subset of deanol-treated patients who had developed tardive dyskinesia and other movement disorders. Thus, precursors of acetylcholine, in addition to cholinergic agonists and anticholinesterase agents, have consistently been noted in several studies to induce a depressed mood. Significant evidence supportive of a role for acetylcholine in the phenomenologyof affective disorders also comes from an analysis of the anergic-inhibitory effects of cholinergic agonists and anticholinesterase agents. These drugs induce a psychomotor retardation which is very similar to that occurring naturally in endogenous depression. This psychomotor retardation includes feelings of fatigue, lack of thoughts, and decreased movements. Thus, Rowntree et al. [1950] and Modestin et al. [1973a,b], studying normals, depressives, and manics, and Gershon and Shaw 119611, observing normals, all reported that cholinesterase inhibitors exerted anergic and behavioral-inhibitory effects. Similarly, Janowsky et al. [197313, 19741 observed psychomotor retardation in their physostigmine-treated subjects, and more recently, Silva et al. 119931 noted anergia, psychomotor retardation, and decreased word generation after physostigmine infusion in a group of carefully screened normal controls. The similarities between the behavioral and other effects of cholinergic stimulation and the symptoms/ correlates of depression are summarized in Table I. Janowsky et al. 11973al noted that the depressive and behavioral inhibitory effects of physostigmine, described above, could be rapidly decreased and antagonized by the noradrenergiddopaminergic drug, intra- zyxwv zyx z Cholinergic Genetics and Affective Illness TABLE I. Similarity of Cholinergically Induced Responses to Symptoms/Correlates of Depression Measure Activity Mood Cortisol ACTH/endorphin Epinephrine REM sleep SW sleep Effects of cholinergic stimulation Depressed symptoms correlates Anergial retardation Depressed Elevated Elevated Elevated Reduced latency None Psychomotor retardation Depressed Elevated Elevated Elevated Reduced latency Reduced amount venous methylphenidate, and vice versa. In addition, methylphenidate, as well as other psychostimulants, closely mimic the symptoms of mania, including euphoria, increased talkativeness and interactions, and insomnia. That study reinforced the possibility that aminergiclcholinergic interactions and balance might be important in the etiology of affective disorders. Differential Effects of Cholinomimetics in Affective Disorder Patients The majority of studies evaluating the effects of cholinomimetic agents on patients with active affective disorder symptoms have demonstrated that these patients are relatively more sensitive to the behavioral effects of cholinomimetics than are controls. With respect to the affect-inducing and behavioral-inhibitory effects of cholinomimetics, Janowsky et al. [1980, 19811 noted that many of those patients described above with symptoms of depression, mania, or schizoaffective disorder, as compared to schizophrenics without a significant mood component in their illness, became significantly more sad and depressed after receiving physostigmine. Furthermore, Janowsky et al. [1980, 19811 found that rater-evaluated increases in behavioral inhibition and self-rated anxiety, depression, hostility, confusion, and decreases in elation subscales of the Profile of Mood States Scale showed significantly greater changes in affective disorder patients than in other psychiatric patient groups or normals after physostigmine infusion. That physostigmine may behaviorally differentiate patients with affective disorder diagnoses from other nonaffective diagnoses has further support from the work of Edelstein et al. [19811. These authors reported that schizophrenic patients who responded to physostigmine with a clearing of psychotic symptoms were significantly more likely to respond positively t o lithium, presumably because they represented a variant of affective disorder. Furthermore, Steinberg et al. [ 19931 noted that increases in negative affect after physostigmine administration occurred selectively in those personality disorder patients with preexisting affectively unstable personalities (i.e., borderline personalities), as compared with those who were affective stable. Thus, it has been demonstrated in several studies that actively ill affective disorder patients exhibit exaggerated behavioral responses t o cholinergic agonists and anticholinesterases, although the degree of differ- 337 ence is not great and the number of replications is small. Conversely, Oppenheimer et al. [1979] and Silva et al. 119931 found no increases in depressed mood in his normal subject cohort, and Rapaport et al. [19921recently reported that panic disorder patients responded similarly to physostigmine as did normal controls, rather than showing the exaggerated responses of depressives. Trait/State Considerations The evidence for behavioral supersensitivity to cholinomimetics as a trait marker in affective disorder patients is mixed. On the positive side, Oppenheimer et al. [1979] observed, in the patients described previously, that a significant percentage of euthymic bipolar patients receiving lithium developed a depressed mood after receiving physostigmine, whereas normal controls who received physostigmine alone did not become depressed, but only became anergic. Similarly, Casey [19791 noted that those euthymic tardive dyskinesia patients with a strong past history of affective disorder selectively showed increased affective symptoms while receiving the putative acetylcholine precursor, deanol. However, Nurnberger et al. [1983al did not observe a differential behavioral sensitivity between a group of euthymic affective disorder patients and normals. Thus, there is presently somewhat conflicting data on whether behavioral supersensitivity in affective disorder patients is a state- or trait-linked phenomenon. Furthermore, no studies published to date have examined the behavioral effects of cholinergic agonists on the unaffected relatives or offspring of affective disorder patients, and thus the stateltrait question regarding cholinomimetic behavioral changes remains an open one. HYPOTHALAlMIC-PITUITARY-ADRENAL AND OTHER ENDOCRINE PARAMETERS z z This section will review lines of evidence suggesting that the hormonal changes exhibited naturalistically by many depressed patients and by normal individuals receiving cholinergic agonists and cholinesterase inhibitors are remarkably similar. In addition, the evidence that hormonal changes exhibited by individuals after the administration of centrally acting cholinomimetics are exaggerated in depressed patients will be reviewed, as will the extent t o which this cholinergically-induced hormonal supersensitivity may be regarded as a state o r trait marker of depression. Phenomenological Similarities Between HPA Changes in Depressives and Central Cholinomimetic Effects In humans, a frequent characteristic of depression, especially severe depression, is the spontaneous activation of the hypothalamic-pituitary-adrenal (HPA) axis. A variety of studies have shown that increased cortisol and adrenocorticotrophic hormone (ACTH) levels are found in patients with endogenous depression, and that cortisol suppression by dexamethasone is inhibited in major depression [Janowsky and Risch, 19841. As with behavioral studies, there is evidence from a variety of 338 zyxwvuts zyxwvuts z Janowsky et al. experiments that centrally acting cholinomimetic drugs release the corticotrophin releasing factor and thus elevate serum ACTH and cortisol levels in a variety of animal species, as well as in normal controls and in psychiatric patients [Risch et al., 1981b,c;Janowsky and Risch, 19841. Also, physostigmine has been shown to reverse dexamethasone-induced suppression of cortisol in normal controls [Carroll et al., 1980; Doerr and Berger, 19831, producing a “depression-like” profile. Thus, it appears that physostigmine and other cholinomimetic drugs cause increases in HPA axis activity, and that these changes parallel other phenomena noted in endogenous depression, such as increased cortisol secretion, cortisol resistance to suppression by dexamethasone, and elevated ACTH levels. In addition, beta endorphin, whose secretion is linked to that of ACTH through CRF induced corelease of a common precursor, beta lipotropin, has also been noted to be elevated in depressed patients, and beta endorphin is released by physostigmine, as well as by arecoline [Risch et al., 1981c, 19821. Differential Effects Between Affective Disorder Patients and Normal Controls Affective disorder patients have been found to show significantly greater increases in both serum ACTH and beta endorphin after physostigmine infusion [Risch et al., 1981b, 19821, when compared to normal controls and t o nonaffective psychiatric patients. Interestingly, this supersensitivity of beta endorphin and ACTH does not appear to occur with respect to cortisol elevation induced by physostigmine or arecoline [Risch et al., 1981b,c, 19821. TraiUState Considerations At present, there are very few studies reporting on the effects of cholinergic stimulation on hormonal responses in ethymic affective disorder patients themselves, or in the healthy relatives of affective disorder patients. Nurnberger et al. [1983al did report a lack of exaggeration of cortisol release following arecoline administration in euthymic bipolar patients, but did not study changes in beta endorphin or ACTH. Consequently, no conclusion about whether the cholinergically-induced supersensitive ACTH and beta endorphin responses in depressives are state or trait markers of affective disorders can be made a t this time. DIFFERENTIAL GROWTH HORMONE RESPONSES TO CHOLINOMIMETICS IN DEPRESSIVES At present, several studies have suggested that growth hormone levels increase when a cholinomimetic drug is given, a phenomena which is blocked by administration of anticholinergic agents [Bruni and Meites, 19781. Possibly due to the presence of anticholinergic drugs given as pretreatments prior to cholinomimetic administration, several groups have failed to find significant increases in growth hormone release after physostigmine andor other cholinomimetic agent infusion in psychiatric patients and in normals pretreated with peripherally active anticholinergic agents such as probanthetine and methscopolamine [Brvni and Meites, 1978; Davis and Davis, 1980; Janowsky and Risch, 1984, 19871. However, recently, OKeane et al. [19921 have reported exaggerated growth hormone release in depressives, especially depressed males, following administration of the peripherally-acting cholinomimetic agent pyridostigmine, when compared to normal controls. In the latter study, none of those evaluated were given a peripherally or centrally active anticholinergic drug prior to cholinomimetic administration. The supersensitive growth hormone response observed in the depressed patients suggests that there could exist peripheral cholinergic supersensitivity in depressives, since pyridostigmine is peripherally acting. This hypersensitivity was not previously apparent, probably due to the use of anticholinergic drugs in the earlier experimental designs. This notion is supported by another recent report indicating that depressives are more sensitive to the effects of pilocarpine on pupillary responses [DeMet and Sokolski, 19931, a finding which, the authors believe, represents a trait phenomenon of depression. It is also well known that growth hormone responses to clonidine, an alpha noradrenergic agonist, are blunted in human depressives [e.g., Charney et al., 1982; Siever and Uhde, 19841. The blunted noradrenergic response and supersensitive cholinergic response could be regarded as support €or the adrenergidcholinergic balance model. However, this blunted growth hormone response t o clonidine is not specific for depressive disorders [Tancer et al., 19931, and there have not been any specific studies in animals or in humans examining cholinergidadrenergic interactions in growth hormone regulation. Further work must occur before we can conclude that the noradrenergic subsensitivity in depressives is in any way connected with the observed cholinergic supersensitivity. SLEEP PARAMETERS This section will review evidence suggesting that the sleep changes which spontaneously occur in depressed patients are very similar to those seen after the administration of cholinomimetic agents, and that the sleep changes induced by cholinergic agonists are exaggerated in individuals with affective disorders. In addition, the extent to which these cholinergically induced sleep alterations are trait markers of the affective disorders will be considered. Phenomenological Similarities Between Sleep in Affective Disorder Patients and the Sleep Altering Effects of Cholinomimetics Although not as diagnostically specific as previously believed, major depression is generally associated with a series of characteristic sleep changes. These include decreased rapid eye movement sleep (REM) latency and increased REM duration and density [Benca et al., 19921. In parallel with these characteristic sleep changes occurring in depression, centrally acting cholinergic agonists and cholinesterase inhibitors, such as arecoline, physostigmine, and pilocarpine induce a shortening of REM latency and an increase in REM du- z zy zyxw Cholinergic Genetics and Affective Illness ration and density [Sitaram et al., 1982, 1985; Berger et al., 1983, 1985, 1989; Berkowitz et al., 19901. Also, Sitaram et al. [19821 have shown that withdrawal of chronic scopolamine also leads to a shortening of REM latency and an increase in REM density. Thus, the REM sleep abnormalities observed in depressed individuals could reasonably be ascribed to cholinergic overactivity, as postulated by Hobson et al. [1975]. Furthermore, in general, adrenergic agents such as psychostimulants and L-DOPA cause the opposite effects of cholinomimetics, shortening REM latency and decreasing REM duration and density. Differential Effects of Cholinomimetic Drugs in Affective Disorder Patients Of all the potential cholinergic markers of affective disorder, changes in REM sleep parameters after cholinomimetic administration have proven the most promising. Sitaram et al. [1987] found that following arecoline infusion, REM latency was shortened significantly more in patients undergoing an affective disorder episode than in controls. Gillin et al. [1991] and Nurnberger et al. [19891 recently replicated Sitaram’s work, showing enhanced cholinergically-induced REM latency shortening in depressives following arecoline infusion. Significantly, Nurnberger et al. [ 19811 also noted an inverse relationship between amphetamineinduced REM shortening and physostigmine-induced REM latency increases, a finding consistent with the adrenergidcholinergic balance hypothesis. In addition, Berger et al. [1989] found that a supershortening of REM latency occurred in endogenous depressives, when compared to normals and to eating disorder patients, following administration of the long-acting oral muscarinic agonist, RS86. Berger et al. [19831 also found that physostigmine-induced arousal and awakening from sleep occurred more frequently in actively ill affective disorder patients than in normals. More recently, Gann et al. [1992] investigated sleep EEG profiles during placebo administration and after cholinergic stimulation with RS86 in patients with major depression, in patients with anxiety disorders, and in healthy control subjects. As in previous studies, RS86 had a more profound impact on patients with major depression, inducing a relative supershortening of REM sleep latency, and an increase in REM density and REM duration. Importantly, patients with anxiety disorders and associated secondary depression did not show enhanced REM abnormalities following RS86 administration. In fact, anxiety disorder patients showed decreased REM density compared to controls. Similarly, Dube et al. [19851 showed that the REM sleep response to cholinergic stimulation with arecoline was significantly more pronounced in primary depressives than in patients with manic disorders, or in those with mixed anxiousldepressive symptoms. These studies support the concept that cholinergic supersensitivity is relatively specific to affective disorders. However, there have been several reports suggesting the involvement of overactivity of REM sleep parameters in the negative symptoms of schizophrenia [Tandon and Greden, 19891, so caution about the ab- 339 solute specificity of cholinergic overactivity in depression must be maintained. Another issue of importance relates back to the adrenergiclcholinergic interaction model described above. While there have been numerous reports of cholinergic supersensitivity in sleep mechanisms in depressives, there have been comparatively few reports regarding other neurotransmitter systems involved in sleep mechanisms. Recently, however, Schittecatte et al. [19921reported that human depressives are subsensitive to the REM sleep-suppressing effects of clonidine, an alpha noradrenergic agonist. This outcome would be predicted from the adrenergiclcholinergiic balance model. A problem is knowing whether the abnormal clonidine response in depressives is an indication of a separate noradrenergic abnormality, or is only reflective of an underlying cholinergic abnormality in balance with the noradrenergic system. It would be valuable to know how these patients would have responded to a cholinergic challenge. Trait/State Considerations There are several lines of evidence suggesting that supersensitive cholinergically-induced REM sleep parameters in depression may have a genetic basis. First, REM sleep responses t o arecoline have been reported to be significantly correlated in normal identical twins [Nurnberger et al., 1983133. Secondly, the work of Sitaram et al. [19871 studying arecoline-induced shortening of REM sleep showed that a supersensitive response was significantly more likely to occur in relatives of endogenous depressives who themselves had a history of affective disorder, than in those relatives who did not have such a history. Even more suggestive of a genetic link between cholinergic supersensitivity sleep parameters is the work of Schreiber et al. [19921. These investigators observed exaggerated shortening of REM latency and increased spontaneous sleep onset REM periods following RS86 administration in healthy first degree relatives of patients with a DSM I11 diagnosis of major depression and a strong family history of emotional disorders. A significantly less extreme response occurred in first degree relatives of individuals without a history of affective disorders. While the above data are supportive of the conclusion that supersensitive REM sleep induction by cholinergic agonists is a trait marker of affective disorders, there are some inconsistent observations. Berger et al. [19891 noted exaggerated REM latency shortening following administration of RS86 only in actively depressed patients, and not in remitted ones. Their findings suggest a state phenomenon. On the other hand, the work of Sitaram et al. [1987] and Nurnberger et al. [1983al would suggest a trait phenomenon, since remitted bipolar patients and individuals with a family history of bipolar disorders all showed exaggerated REM latency shortening after receiving arecoline. One of the major differences between the latter studies and those of Berger et al. [1989] was the fact that the large majority of Berger et a1.k patients were diagnosed as having unipolar depression, as compared to the bipolars studied in the investigations of Sitaram et al. [19871 and 340 zyxwvutsrqp zyxwvuts Janowsky et al. Nurnberger et al. [1983al. Thus, cholinergic overactivity may indeed be a trait marker for bipolar affective disorder, and a state marker for unipolar affective disorder. This conclusion would fit well with the evidence from epidemiological studies for a stronger genetic link in bipolar affective disorders [Gershon et al., 19871. CENTRAL MUSCARINIC MECHANISMS MIMICKING DEPRESSION Several studies have attempted to define the mechanisms by which the behavioral, sleep, and neuroendocrine-altering effects of cholinomimetic drugs occur. In early studies, Janowsky et al. 119721 and Modestin et al. [1973a,bl noted that, in contrast to centrally acting physostigmine, the peripherally acting anticholinesterase, neostigmine, did not exert any behavioral, neuroendocrine, or cardiovascular effects if a peripheral anticholinergic had been used as a pretreatment. This observation indicates that the behavioral effects of physostigmine were probably due to its central actions. More recently, Janowsky et al. [ 19851 noted that the increases in blood pressure, pulse rate, serum epinephrine, ACTH, cortisol, and prolactin, as well as the anergic and negative affects induced by physostigmine, were not mimicked by neostigmine, again supporting a central mechanism as the basis for cholinomimetic-induced changes. Furthermore, Janowsky et al. [19851 noted that the behavioral, cardiovascular, and neuroendocrine effects of physostigmine could be blocked by the centrally acting antimuscarinic agent, scopolamine, but not by the peripherally acting antimuscarinic agent, methscopolamine. These findings again indicate the involvement of central cholinergic mechanisms in the behavioral, neuroendocrine, sleep and cardiovascular effects of cholinomimetic agents, and, in addition, indicate that muscarinic cholinergic receptors are involved. The exact location of the cholinergic subsystems in the brain which mediate the exaggerated muscarinic responses in depressives described above is unknown. Meyerson et al. [1982] reported preliminary evidence for an elevation of cortical muscarinic receptor binding sites in brains from suicide victims, presumed to have affective disorders. However, both Stanley 119831 and Kaufman et al. 119843failed to detect any differences in cortical muscarinic receptor binding between suicide victims and controls. Studies which have examined muscarinic receptors on fibroblast cells in affective disorder patients have also ultimately been negative [Nadi et al., 1984; Kelsoe et al., 19851. A problem with all of the studies performed to date may be that the tissues selected for study may have been from inappropriate locations. There are data that the Flinders Sensitive Line (FSL) rats, bred t o have exaggerated sensitivity to cholinergic drugs, do not necessarily have elevated muscarinic receptors in all brain regions, and, indeed, do not exhibit elevated cortical muscarinic receptor binding sites [Overstreet, 1993; see below]. It would be of interest to obtain muscarinic receptor binding data from limbic regions in normal and depressed humans, because the cholinergically supersensitive FSL rats have been found to have elevated limbic muscarinic receptor binding sites [Overstreet, 19931. At present, however, it is not possible to ascribe the supersensitive muscarinic responses in depressives to any specific brain region, or to an excess of central muscarinic receptors. There have also been a limited number of studies which have searched for a linkage between muscarinic receptor genes (five have been identified to date) and affective disorders. These studies have also been inconclusive [Berrettini et al., 1992; Nurnberger et al., unpublished data]. Therefore, while it seems clear that the supersensitive muscarinic receptor responses in depressives are centrally mediated, it is not clear which region(s) of the brain is (are) involved, nor is it known whether muscarinic receptor genes are linked t o affective disorders. Also, it may be that supersensitive muscarinic responses are mediated not by muscarinic receptors per se, but by one or more of the many second messenger systems that are activated via muscarinic mechanisms such as the G proteins [Lesch and Manji, 19921, or by a decrease in adrenergic neurotransmitters. zyxwvu ACETYLCHOLINE, STRESS, AND DEPRESSION Considerable information suggests that a propensity to stressability may be a hallmark of depression, and that indeed depression may be a specialized form of stress. Janowsky and Risch [1984] have reviewed evidence suggesting that most of the manifestations of stress in general (i.e., behavioral, cardiovascular, and neuroendocrine measures), which also occur in endogenous depression, are replicated by central cholinergic activation. These cholinomimetic stress-like effects are generally exaggerated in depressives. Similarly, Overstreet 119931 has noted that FSL rats, selectively bred to be sensitive to cholinomimetics, are hyperresponsive to stress, show a mimicking of depression-analgous behaviors, and are supersensitive to cholinomimetic drugs with respect to stress-indicative phenomena. Finally, there is considerable evidence that central acetylcholine activates CRF release, leading to the neuroendocrine cascade indicative of stress. Thus, it is quite possible that a genetic defect in the modulation of acetylcholine output or receptivity could lead to a series of exaggerated responses to stress, one of which could be depression. zyx NICOTINE, SMOKING, AND DEPRESSION Although the vast majority of observations linking the cholinergic nervous system to affective disorders has focused intensively on muscarinic mechanisms, there is also evidence that nicotinic cholinergic mechanisms may be linked to depression. With respect to nicotine’s effects in humans, Glassman 119931 has extensively reviewed his own work and the work of others indicating that a high rate of cigarette smoking is associated with current major depression and current depressive symptoms. He also noted that a lifetime history of major depression, even if not active a t the time that a person starts t o smoke, increases the chances of a person’s trying nicotine and becoming addicted to it, and has a significant negative impact on smoking cessation efforts [Glassman, 19931. This latter deleterious z zy zyxwv Cholinergic Genetics and Affective Illness effect appears more pronounced in women than in men. There is also evidence that in predisposed individuals with a history of major depression, smoking cessation may precipitate severe depressive symptoms which appear to counteract smoking cessation efforts. Of course, the linkage between cigarette addiction, smoking, and emotional disorders is not exclusive for depression. There are other linkages between smoking and alcoholism, anxiety disorders, and especially schizophrenia [Glassman, 19931. Another interesting linkage between smoking and depression has been noted in twin studies. Using monozygotic and dizygotic twin pairs, Kendler et al. [ 19931have found that smoking and depression are indeed linked, but that smoking, at least in their study, did not necessarily cause depression, and that depression did not necessarily cause smoking. It appears that depression and smoking are linked through genetic factors which influence vulnerability to both conditions [Kendler et al., 19931. It is possible that a relationship between muscarinic and nicotinic mechanisms may be important in inducing nicotine addiction, in inducing depression, and in the relationship between smoking and depression noted above. Possibly, an interaction between muscarinic and nicotinic receptors may underlie this relationship. 341 other rat strains which appear to be anergic a t baseline have also been reported to be more sensitive to cholinergic agonists [Overstreet et al., 1988, 1992al. Recently, Overstreet et al. [1992b] crossbred FSL and control Flinders Resistant Line (FRL) rats and studied their muscarinic responses. These investigators found that both additive and dominance genetic variance contributed to many of the responses of the F2rats before and after cholinomimetic administration. They estimated that 3-15 genes might be involved. In an even more recent study, both immobility in the swim test and hypothermic responses to the cholinergic agonist, oxotremorine, were examined in the FSL/FRL crossbreeds. Again, it was found that the degree of oxotremorine-induced hypothermia was dependent on both additive and dominance genetic factors, with the crosses more closely resembling the FRL control parent [Overstreet et al., 1993a,b]. However, swim test immobility appeared to be dependent solely upon additive factors [Overstreet et al., 1993a,bl. This finding is consistent with one of two models. In the first, the genes for immobility and muscarinic sensitivity are overlapping, but have different outcomes for the two measures, similar to albino genes exhibiting dominance for coat color and additivity for enzymes. In the second model, the genes for immobility and muscarinic sensitivity are independent. Further studies with appropriate genetic animal ANIMAL ANALOGS AND PARALLELS models can provide further definition and possibly Administration of cholinergic agonists to animals point to similar phenomena in humans. For example, produces a series of behavioral and physiological phe- recombinant inbred strains of mice may be utilized to nomena that parallel those seen in humans. These in- locate single genes important in behavioral or neuroclude anergic-inhibitory effects, anhedonic phenomena, chemical variation [Gora-Maslak et al., 19911. Nurnhypothalamic-pituitary-adrenalactivation, shortening berger et al. [in press] report that the C57BU6J strain of REM sleep latency, and the exaggeration of REM of mouse shows increased choline uptake in the striasleep density. In addition, as with humans, the anergic tum, compared to the DBA/2J strain [cf. Charles et al., effects of cholinomimetics in rodents can be reversed by 19931. The C57s are also more sensitive to the forced psychostimulant (i.e., methylphenidate) administra- swim and restraint stress models of depression. Quantion; and methylphenidate-induced stereotyped gnaw- titative trait loci for these parameters have now been ing behavior (a dopaminergic phenomena) can be re- identified using recombinant inbred strains [Tarricone versed by physostigmine [Janowsky et al., 19721. et al., 19931. The Flinders Sensitive Line (FSL) rats are an AusCONCLUSION tralian line of rats selectively bred to have increased responses to anticholinesterases and cholinergic agonists As mentioned above, a number of studies suggests [Overstreet et al., 1979,19881. These rats resemble de- that patients with an affective disorder show a tenpressed humans in that they exhibit supersensitive be- dency to have either exaggerated reactions to centrally havioral, hormonal, and sleep responses to cholinergic acting cholinomimetic agents or an excess of central agonists relative t o their control counterparts, the acetylcholine. This supersensitivity appears to occur Flinders Resistant Line (FRL) rats [Overstreet et al., more often in patients with endogenous depression or 1988; Overstreet, 19931. The FSL rats also resemble bipolar disorder. The data are most compelling with redepressed humans in that at baseline they show de- spect to supersensitive responses of cholinomimeticcreased running activity [Overstreet and Russell, sensitive sleep parameters following cholinomimetic 19821, increased anhedonia (i.e., reduced saccharin administration in affective disorder patients, especially consumption) under stress [Pucilowski et al., 19831, with regard to REM sleep measures. At least one study and increased REM activity and density [Shiromani et of high risk members of families loaded for affective disal., 19881. Thus, a line of rats established to have orders suggests that this supersensitivity might be a cholinergic hyperactivityfiypersensitivity resembles trait phenomenon [Schreiber et al., 19921, and other depressed humans in a number of ways. Moreover, studies by Nurnberger et al. [1983al and Sitaram et al. anergic-like effects in the FSL rats (i.e., exaggerated [19871 are similarly supportive. immobility in the forced swim test) can be counteracted Studies showing a differential induction of depressive by the administration of antidepressant drugs [Schiller symptoms in euthymic individuals with a history of afet al., 1992; Pucilowski and Overstreet, 19931. Several fective disorders have not been as consistent. In addi- zyxwv 342 zyxwvutsrqp zyxwvuts Janowsky et al. tion, studies of the behavioral effects of cholinomimetics in high risk populations have not yet been carried out, nor have studies of neuroendocrine, biochemical, and physiological markers, including brain choline levels, cholinomimetic-induced increases in ACTH and beta endorphin, and decreases in pupillary size. To date, cholinomimetic agonist and cholinesterase inhibitor strategies have primarily been used to explore the adrenergiclcholinergic hypothesis of the etiology of affective disorders and the role of cholinergic super-sensitivity in depression. Only a few studies have explored the psychogenetic aspects of cholinergic hyperresponsiveness or adrenergic-cholinergic balance as related to affective disorders. Nevertheless, as described above, there is a growing body of information suggesting that the cholinergic hypersensitivity found in affective disorder patients may indeed be genetically determined, and, thus, cholinergic hyperresponsiveness may be a reasonable candidate for marking affectively vulnerable members of families. Targeting genes which influence or regulate cholinergic function may reveal candidate genes which can be linked to vulnerability in families with high levels of affective disorder. To expand upon the above point, a number of future genetically-oriented studies and strategies exploring the role of acetylcholine and adrenergic cholinergic balance in the affective disorders would be appropriate. Expansion of the rodent crossgenerational studies and crossbreeding studies, described above, with an expansion of acetylcholine sensitive variables, is indicated. Such studies, using quantitative locus analysis, may help t o characterize cholinergic regulatory genes and their behavioral correlates. Recently, Hwang et al. [in preparation] used the recombinant inbred strain method to localize a gene responsible for regulating choline uptake in the striatum, hippocampus, and cortex. Similarly, studies should be designed to locate and characterize candidate cholinergic genes in man, including muscarinic receptor genes, genes which regulate choline uptake and utilization, and genes which regulate choline acetyltransferase activity. Linkage and association studies in families with affective disorder are appropriate, focusing on these and other candidate genes. Human studies should also include imaging experiments using presynaptic cholinergic agents or agents that label cholinergic receptors. Finally, it is appropriate to speculate as to why a genetic tendency to have a hyperreactive cholinergic nervous system might be of survival value. One possibility is that cholinergic hyperactivity is involved in the regulation of the hypothermia, inactivitylanergia, and sleep changes of hibernation, or in the “giving up” responses of animals confronted and overpowered by predators andlor dominant members of their own species. 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