Exploring the brain gut axis in irritable bowel syndrome specific emphasis on stress and melatonin

Symptoms, such as abdominal pain and altered motility and bowel habits in IBS patients can arise from dysregulation of activity in one or more of the stations in the bidirectional commun

Part I

Literature Review and Hypothesis

Chapter 1

Spectrum of Irritable Bowel

Syndrome and Brain-Gut Axis

1.1 Introduction

The irritable bowel syndrome (IBS) is a common disorder producing abdominal pain, bloating and altered bowel habits, which can be associated with significant disability and health care costs IBS is defined by symptoms, which are markedly influenced by psychological factors and stressful life situations, in the absence of structural pathology No life threatening or significant disease such as infections, inflammatory bowel disease or bowel cancer have been found in majority of IBS patients on appropriate investigations With the accumulation of knowledge from epidemiology, physiology, psychology and neuroscience during the last decade, IBS

is now believed to result from a dysregulation of brain-gut axis which involves the alterations in intestinal motor, sensory, and central nervous system (CNS)

The histories of patients with IBS show considerable variation in symptom experience and behavior Though most of these patients only have relatively mild to moderate symptoms, some suffer severe symptoms that can restrict social activities and substantially reduce quality of life and cause serious consequence It can affect employment, leisure travel, sexual function, diet and co-morbid with sleep, depression and anxiety Patients with IBS have three times work absenteeism than other employees (Drossman et al, 1993), and based on the large proportion of the population affected, IBS patients consume tremendous healthcare services IBS accounts for an estimated 2.4-3.5 million physician visits per year in the United States (Everhart and Renault, 1991) and for an estimated 2.2 million medication prescriptions (Sandler, 1990) Patients with IBS undergo numerous diagnostic tests

and procedures (many unnecessary), and may retain a maladaptive illness belief that some other diagnosis is still being missed Some of them are also more likely to undergo surgery (Longstreth et al, 1990)

The cost of health services for patients with IBS is significantly higher than that for controls and is estimated at eight billion dollars per year for the white population of the United States (Talley et al, 1995) Because of poor understanding of the cause of the disorder, lack of common-accepted diagnostic criteria and a biopsychosocial model of the disease, there is still no encouraging strategy for cure and the optimal treatment for IBS remains controversial As such, we face many problems and challenges and there is clearly a need for more research on this disorder

1.2 Epidemiology

In general, large epidemiological studies from United States, United Kingdom, and China show that IBS affects about 11%-20% of people in the community (Farthing, 1995) The prevalence of IBS varies across different epidemiological studies; presumably due to the diversity of definitional criteria, differences in the specific questions used to elicit the information, different target population and other factors

A survey using Manning criteria found that 22% of the British population aged between 20 and 90 years have IBS (Jones and Lydeard, 1992) Another study using the same diagnostic criteria but in a different age group (30 to 64 years), reported a prevalence of approximately 17% (Talley et al, 1991) Using the more restrictive Rome criteria, IBS symptoms was only detected in 9.4% of the United States population (Drossman et al, 1993)

It has also been documented that only a proportion of those with IBS symptoms consult physicians While a study done by Talley et al (1991) reported that only 14%

of patients with IBS symptoms had consulted a physician, a survey conducted by Jones and Lydeard (1992) revealed that only one-third IBS patients had seen a doctor for their symptoms

There are marked differences in the prevalence of IBS in women as compared with men IBS affects females approximately twice as often as males In the U.S Householder Survey (Drossman et al, 1993), IBS was present in 14.5% of women but

in only 7.7% of men Similar differences in prevalence have been reported in other studies from western countries (Hislop, 1971; Talley et al, 1991; Heaton et al, 1992; Jones and Lydeard, 1992) Contrary to findings by the abovementioned studies done

in the western countries, studies from India and Sri lanka reported a preponderance of men having IBS (Kapoor et al,1985; Mendis et al, 1982) The disparity of IBS prevalence between the western and Indian reports may be attributed to different healthcare seeking tendency between men and women in the two different societies (Thompson et al, 1989)

1.3 Etiology and pathophysiology

The pathophysiology of IBS remains largely unknown Several mechanisms have been postulated as the basis for the cause and development of IBS These mechanisms include dysregulation of brain-gut interactions, psychological factors, abnormal motility, enhanced visceral sensitivity and autonomic system imbalance However, no mechanism unique to IBS has been identified

1.3.1 The brain-gut axis

Currently, more and more attention has been put on the dysregulation of brain-gut interactions The brain-gut axis refers to the continuous back and forth interactions of information and feedback that take place between the gastrointestinal tract, and the brain and spinal cord These interrelated feedback circuits can influence brain processes and bowel functions including pain perception, gut sensitivity, secretions, inflammatory responses, and motility The brain-gut circuits can be activated by an external or internal factor or stimulus that makes a demand on the system, such as a stressful event Symptoms, such as abdominal pain and altered motility and bowel habits in IBS patients can arise from dysregulation of activity in one or more of the stations in the bidirectional communication pathways between the GI system (the enteric nervous system, ENS) and the spinal cord and brain (the central nervous system, CNS)

Through the use of modern imaging techniques, such as positron emission tomography (PET) and functional magnetic resonance imaging (fMRI), researchers have been able to evaluate cerebral blood flow or oxygen desaturation respectively, in areas of active brain functioning in response to real or anticipated pain from visceral balloon distention A fMRI study found increased anterior cingulated cortex (ACC) activation in IBS patients compared to healthy controls as well as activation of prefrontal, insular and thalamus in most subjects in response to painful rectal distention (Mertz et al, 2000) However, using PET, it was also revealed that unlike healthy controls, IBS patients failed to activate the ACC during painful rectal

distention (Silverman et al, 1997) Such differences might suggest these findings are related to anxiety and uncertainty among the patients In spite of these inconsistent results, these data at least provided an objective evidence to support the hypothesis of the dysregulation of brain-gut communication, and such dysregulation are especially manifested in the abnormal descending pain inhibitory modulation

Numerous neurotransmitters found in the brain and gut act as messengers that regulate brain-gut communication under stress These messengers, including serotonin (5-HT), cholecystokinin, substance P, enkephalins, calcitonin gene related polypeptide, nitric oxide, and others, have varied and integrated effects on pain modulation, gastrointestinal motility, emotional behavior, and immunity (Mayer and Gebhart, 1994) Given the complex relationship between inflammatory mediators, gut hypersensitivity, motility, and pain experience, the results of recent research strongly suggest that alterations in neuroimmune and neuroendocrine communications at the enteric and CNS levels may trigger a series of events that gives rise to chronic changes in visceral sensitivity and central mechanisms controlling pain, as evidence

of dysregulation of the brain-gut axis Several kinds of events could trigger the dysregulation of the brain-gut axis in patients with IBS Among these are psychological experiences, such as life stress, psychological co-morbidity, or sexual and physical abuse, and inflammation (Drossman et al, 1996; Gwee et al, 1999)

1.3.2 Stress

Stress, defined as an acute threat to the homeostasis of an organism by real (physical, ‘interoceptive’; e.g gut infection, cold water immersion, visceral distention)

or perceived (psychological, ‘exteroceptive’; e.g dichotomous listening, mental arithmetic, life events) events, initiates adaptive physiologic and behavioral changes that serve to defend the stability of the internal environment (Monnikes et al, 2001; Selye,1998; Selye, 1976; Chrousos and Gold, 1992) Certain physical and psychological stresses have been associated with the onset or symptom exacerbation

in functional gastrointestinal disorders (FGD), especially IBS Such association in patients with IBS was observed in some well-designed surveys (Gwee et al, 1999; Bennett and Tennant et al, 1998; Whitehead et al, 1992; Welgan et al 1985) Patients with FGD are exposed to one or more stressors much more often than normal controls (98% versus 36%) (Bennett and Piesse et al, 1998) Meanwhile, the symptoms are improved with the acquisition of more effective stress management skills (Guthrie et

al, 1993; Shaw et al, 1991) Furthermore, evidences from animal studies have shown that various stressors caused delayed gastric emptying, inhibited small bowel transit and accelerated colonic transit (Williams et al, 1988; Tache et al, 1999; Tsukada et al, 2002) Some investigators also observed increased responses of distal colonic motility

in response to stress in IBS patients (Welgan et al, 1988; Fukudo et al, 1993; Narducci et al, 1985)

Recently, a study group reported decreased rectal pain thresholds in patients with IBS but not in normal controls during acute laboratorial physical (cold water hand immersion) and psychological (dichotomous listening) stress (Murray et al, 2004) By contrast, a variety of stressful stimuli has been shown to produce analgesia, a phenomenon often referred to as stress-induced analgesia (SIA) (Bodnar, 1986; Hayes and Katayama, 1986; Watkins et al, 1982) Studies using different stressors

(e.g cold stress pain, ischemic pain and noxious heat) significantly produced analgesia in healthy subjects (Washington et al, 2000; Willer et al, 1989; Pertovaara

et al, 1982) However, this “stress inhibiting pain” phenomenon has never been reported in patients with IBS Taken together, all these data may imply an altered stress regulatory mechanism in IBS patients, especially endogenous pain modulation, which change the sensory and motility of the bowel These changes in turn could cause visceral hyperalgesia, abdominal symptoms and altered stool habits

Compared with the knowledge in physiology, the role of central functions in stress and thereby the interactions between neural networks and gut remains poorly understood Sawchenko et al pointed out that there are some similar principal circuits underlying the stress response in spite of the different types of stress (Sawchenko and

Li, 2000) The limbic forebrain including the lateral and medial prefrontal cortex, hippocampus, and amygdale may participate in central processing psychological stress (Sawchenko and Li, 2000) A recent study observed significant group differences in the frontal brain among healthy controls and IBS patients with respect

to the event-related potentials in the brain during exposure to everyday words with emotional content (Blomhoff et al, 2000) Using electroencephalograms (EEG), Nomura et al also showed significantly greater EEG abnormality in the IBS patients (29.2%) than in the controls (4.2%) under mental arithmetic stress and the administration of neostigmine (Nomura et al, 1999)

All these data suggest a possible involvement of the CNS in the pathophysiology of IBS, especially under the stress situation

1.3.3 Psychopathology

Some chronic and acute psychosocial factors including early life experiences, conditioning factors (Levy et al, 2000; Whitehead et al, 1994), psychological stress (which has been introduced in the section 1.3.1), personal and social coping systems (Drossman et al, 2000), and psychological distress and co-morbidity, are important in the pathophysiology of IBS A whole variety of chronic psychopathologies have been described in secondary care IBS patients although anxiety and depression are by far the commonest, accounting for at least 60% (Whitehead and Crowell, 1991) of them Abnormal illness behavior and illness attitude have also been reported as being more common in IBS than healthy controls (Drossman et al, 1988; Levy et al, 2000) Compared with healthy controls, patients with IBS are observed to have higher scores for anxiety, depression, hostile feelings, sadness, interpersonal sensitivity, as well as more sleep disturbance (Whitehead et al, 1980; Svedlund et al, 1985; Gomborone et

al, 1995; Ford et al, 1987) All of these factors could cause chronic stress in IBS subjects This opinion was furthere supported by the finding that many patients with IBS have counterproductive coping styles, such as cognitions that "catastrophize" symptoms and life events (Drossman et al, 2000) In another study, psychological factors were reported to predict the development IBS after an episode of acute gastroenteritis in previously asymptomatic individuals (Gwee et al, 1999) The prevalence of psychiatric diagnoses in IBS ranges between 40% and 100% depending

on the population, settings, and diagnostic criteria (Hochstrasser and Angst, 1996; Drossman et al, 2000) However, part of these findings maybe overestimated because the data are drawn from patients who are selectively referred to medical centers It

was found that persons with IBS who do not see physicians are psychologically similar to normal subjects, and conversely, frequent clinic attenders have a high frequency of psychosocial disturbance (Drossman et al, 1997; Drossman et al, 1988; Smith et al, 1990) Hence, psychosocial factors not only influence the individual’s illness seeking behavior but also may affect the way in which symptoms are perceived and evaluated

1.3.4 Altered motility

Abnormal bowel motility has been found in patients with IBS since long time ago (Harding, 1951), and IBS has also been long considered as a motility disorder Early studies suggested abnormal electrical control activity in the colon of IBS patients (Snape, 1976) Compared to healthy controls, both a reduction in and an increase in the number of contractions per minute were found in IBS patients (Snape, 1976) Accelerated transit in the colon was observed in some diarrhea predominant IBS patient; on the other hand, delayed transit was found in some constipation predominant IBS patients (Cann et al, 1983; Bassotti et al, 1988) Recently, a study suggested that IBS patients have impaired transit and tolerance to intestinal gas and this anomaly has been cited as a possible mechanism for pain and bloating which are commonly observed in IBS patients (Serra et al, 2001) Furthermore, abnormal motility patterns have also been found in other regions of the gastrointestinal tract, such as the small bowel in IBS patients (Kellow and Phillips, 1987) The transit time through the small bowel was observed to be accelerated in diarrhea predominant IBS patient, but delayed in constipation predominant IBS patients (Cann et al, 1983)

However, the types of motility patterns seen in the colon and small intestine of IBS patients are qualitatively similar to the contractions seen in healthy controls Most stool weights in IBS patients lie within the normal range in spite of their fast transit, and this phenomenon was even found in diarrhea predominant IBS patients (Arffmann et al, 1985; Cann et al, 1984; Vassallo et al, 1992)

In addition to the differences between IBS patients and healthy controls in the basal motility pattern, patients with IBS have exaggerated intestinal motor in response to environmental or enteric stimuli Colonic activity is significantly accelerated in IBS patients when they experience psychological stress (kendall et al,1990; Fukudo and Suzuki, 1987) Eating a meal stimulates colonic activity, and gastrocolonic reflex is prolonged and exaggerated in IBS (Sullivan et al, 1978; Rogers et al, 1989) Moreover, these exaggerated responses to eating are found to be associated with abdominal pain (Sullivan et al, 1978) Rectal distention studies also show a greater increase in colonic motor activity in IBS patients compared to healthy controls (Whitehead et al, 1990) Pharmacologic stimulation of the bowel with cholecystokinin generated more contractile response in IBS patients than in healthy controls and this exaggerated response was shown to persist (Harvey and Read, 1973)

In summary, although IBS patients have motility abnormalities, none of them are specific for IBS In fact, many abnormal motor patterns observed in IBS patients are also found in healthy subjects (McKee and Quigley, 1993; Cann et al, 1983) Thus abnormal motor patterns are not diagnostic for IBS (McKee and Quigley, 1993; Drossman et al, 1997) Furthermore, these abnormal motility patterns neither correlate with IBS symptoms nor are they sufficient to explain many of the symptoms

associated with IBS Therefore, nowadays, abnormal motility is usually no longer considered the primary cause of IBS and it is generally believed that other mechanisms may also be involved

1.3.5 Visceral hypersensitivity

The studies in rectosigmoid balloon distention and the above mentioned specific gut motility changes in IBS patients shifted the attention of researchers to alterations in visceral sensations, i.e visceral hypersensitivity in the 1970s The finding that IBS patients have lower pain thresholds was first described in rectal and sigmoid colon using rectosigmoid balloon distention test (Ritchie, 1973) Thereafter, similar results were also found elsewhere of the gastrointestinal tract including the descending colon, small intestine (Kellow and Phillips, 1987), stomach and esophagus (Trimble et al, 1995) Other than these gastrointestinal tract hypersensitivity, the complain of extra-intestinal manifestations such as headaches, breathlessness, chest pain, fatigue, dysuria and dyspaneuria were often found in IBS patients (Whorwell et al, 1986; Talley et al, 1991; Jones and Lydeard, 1992) All these data may suggest that IBS patients may either have a lowered sensory threshold

non-or have a tendency of apprehension to pain and other symptoms This hypersensitivity

is specific to visceral stimulation because somatic pain thresholds to extreme cold or transcutaneous electrical stimulation are not lower in IBS patients when compared to healthy controls (Whitehead et al, 1990; Cook et al, 1987) However, a response bias should not be omitted, because IBS patients are much more likely to show an

increased hypersensitivity when the rectum is distended in a predictable increasing volume sequence than in a random iterative method

IBS patients may be prone to developing sensitization because repetitive painful balloon distention in the sigmoid colon induced hypersensitivity in the rectum of IBS patients but not healthy controls (Munakata et al, 1997) Such sensitization seems to occur at the level of the lumbar dorsal horn neurons because the dorsal horn neurons

in the spinal cord receive converging signals from the sigmoid colon and rectal splanchnic afferents

In summary, the pathophysiological mechanism of IBS remains to be clearly elucidated However, an integrated model which attempts to synthesize the factors discussed above has been pointed out (Figure 1-1) (Drossman, 1998; Ringel et al, 2001) In this model, IBS is neither a simple physiological nor a simple psychological illness It results from a dysregulation of brain-gut interactions that affect intestinal as well as central and peripheral neurological function, mediated through neuroendocrine and neuroimmunological pathways Psychosocial factors do not cause IBS but influence the expression of the symptoms and illness behavior (Ringel et al, review, 2001)

Figure 1-1 Conceptual model for irritable bowel syndrome (Adapted from Ringel et

al, 2001)

1.4 Diagnosis

A correct diagnosis of IBS is of crucial importance because it reassures patients about the favorable prognosis of their disease and places the basis for a therapeutic strategy on controlling the predominant symptom (e.g constipation, diarrhea, alternating bowel and pain/bloating) The diagnosis of IBS is based on recognizing clinical symptoms consistent with it and on modest efforts to exclude other/organic diseases, because there are no positive physical findings or biological markers for IBS (Drossman et al, 1997; Ringel et al, 2001)

In 1978, Manning et al described several key abdominal symptoms, now referred to

as 'Manning criteria', that were more likely to be present in the IBS than in organic abdominal disease (Manning et al, 1978) (Table 1-1) Kruis et al added other criteria, including a requirement for symptoms to have been present for more than two years and the use of symptom complexes that increase the chances of making a positive clinical diagnosis (Kruis et al, 1984) These symptom criteria were integrated later into other scoring systems The Rome I, and more recently, the Rome II criteria are the results of multinational ad hoc consensus workshops (Thompson et al, 1999) (Table 1-1) Notably, factor analysis data proved that IBS can be defined by a cluster

of three symptoms which represent the core of Rome II criteria Although the Rome criteria are becoming more popular and have been accepted by many regulatory authorities, the Manning criteria are better validated Both Rome I and II criteria proved to be valuable in identifying similar proportions of IBS patients in the general population; however, Rome I criteria allowed the identification of higher numbers of

IBS patients in referral centers than Rome II It is not known why Rome I and II work similarly in the community but yield different results in referral centers It is probable that inappropriate use and/or intrinsic limitations of diagnostic criteria may be responsible for this discrepancy Overall, the general view is that the Rome II criteria are extremely valuable as research tool rather than useful criteria in daily practice (De Giorgio et al, review, 2004) Furthermore, table 1-2 summarizes the main studies that should be considered according to the patient's predominant symptom

Table 1-1 Symptom-based criteria so far established for the diagnosis of IBS

Manning

Pain relieved by defecation

More frequent stools at the onset of pain

Looser stools at the onset of pain

Visible abdominal distension

relieved with defecation

associated with change in frequency of stools

Associated with change in form of stools and two or more of the following

symptoms:

altered stool frequency and/or form

altered stool passage

relieved with defecation

associated with change in frequency of stools

Associated with change in form of stools with the following symptoms

supporting IBS:

altered stool frequency and/or form

altered stool passage

passage of mucus

bloating or abdominal distension

TABLE 1-2 Further evaluation of irritable bowel syndrome optional studies

(Adapted with permission from Ringel et al, 2001)

Constipation-predominant IBS

Colonic transit time (e.g Sitzmark study)

Anorectal manometry and rectal balloon expulsion

Electromyography

Defecography

Diarrhea-predominant IBS

Stool tests (e.g volume, electrolytes, fat, white count, osmotic gap, and pH)

Breath tests for bacterial overgrowth

Intestine absorption function tests (e.g D-xylose)

Small bowel radiograph (partial obstruction, inflammatory bowel disease)

Upper endoscopy with duodenal aspirate (Giardia) and biopsy (celiac sprue,

Whipple's, and lymphoma)

Colonic biopsies (collagenous/lymphocytic colitis)

Serum hormones/peptides (i.e Gastrin, VIP, 5HIAA)

Stool and urine analysis for laxative

Discomfort/pain-predominant IBS

Upright abdominal X-ray (chronic pancreatitis and chronic idiopathic pseudo- obstruction)

Small bowel radiograph (partial obstruction, inflammatory bowel disease)

Additional abdominal imaging studies (i.e X-ray, ultrasound, computerized

tomography, magnetic resonance)

Serum amylase (chronic pancreatitis)

1.5 Treatment

The treatment of IBS involves both pharmacological and non-pharmacological approaches, the latter of whichappear to have the best long term results, particularly

in terms of dietary and psychotherapy treatments

1.5.1 Non-pharmacological treatment approach

1.5.1.1 Construction of good therapeutic relationship

Once the diagnosis of IBS is established, an effective physician patient relationship is crucial for setting the basis of a beneficial pharmacological approach This includes effective communication, appropriate reassurance, and patient education about the condition and its consequences (Drossman et al, 1999) Physicians should explain to patients the nature of their condition, treatment options, and impact of anxiety and stress on symptoms Evidence indicates that a positive interaction with patients with discussion of precipitating factors, diagnosis, and treatment has been shown to reduce the number of returnvisits (Owens et al, 1995)

1.5.1.2 Life style and dietary modification

Many IBS patients benefit from dietary changes, lifestyle changes, and promoting behaviors Some patients with mild IBS symptoms may not require prescription medications after appropriate changes in life style and diet Identifying offending dietary substances (e.g lactose, caffeine, fatty foods, alcohol, gas-producing foods, sorbitol) and making dietary modifications can help some patients

health-An increased dietary fiber intake is one of the most common recommendations in

constipation-predominant IBS Potentially, fibers may be beneficial in constipated IBS patients because of their ability to reduce the transit time through the entire alimentary tract and intestinal wall tension by decreasing intracolonic pressure (Muller-Lissner, 1988) However, physicians should keep in mind that patients may get excessive on dietary restriction which may turn out to be potentially dangerous for them Psychological stressors, which are known to maintain and exacerbate symptoms, should be sought and managed with supportive advice or lifestyle modification Simple life style modifications such as exercise and defecating patterns may help individual patients

1.5.1.3 Psychological treatment

Psychological treatment, including cognitive behavioral therapy, psychotherapy, relaxation and hypnosis, may represent a valid therapeutic option, at least in a sub-group of patients with IBS A small relaxation therapy showed a reduction in symptoms and consultations (Jones et al, 2000) Biofeedback in conjunction with relaxation therapies has also shown some benefit (Lynch and Zamble, 1989; Neff and Blanchard, 1987), but these study results may be biased by the effects of good relationship between therapist and patients due to methodology shortcomings The role of cognitive behavioral therapy and hypnotherapy has not been clearly defined, although small studies suggest some response with symptom improvement (Whorell, 1987; Payne and Blanchard, 1995) But another randomized trial showed hypnotherapy as one of the most valuable non-pharmacological treatment in IBS, with benefit lasting longer than 12 months (Francis and Houghton, 1996).It should be

recognized that the availability of these psychological therapies is limited and their use should be restricted to certain most difficult cases

1.5.2 Pharmacological treatment

The multiple symptoms that are so characteristic of IBS have proven to be a hurdle

in the treatment of the condition Current pharmacological treatment of IBS is only aimed at controlling the predominant symptoms: abdominal pain/bloating,diarrhoea,

or constipation (Camilleri et al, 2002); therefore, it has limited efficacy overall (Camilleri and Choi, 1999) Patients should be informed that any prescribed compound should be taken only during symptom recurrence rather than chronically

1.5.2.1 Abdominal pain and bloating

The most common medications used for management of pain in IBS patients are the antispasmodics, which include three major subclasses, namely anti-muscarinics smooth muscle relaxants and calcium channel blockers (De Giorgio et al, 2004) The final effect of all these compounds is to evoke smooth muscle relaxation and hence reduction of gut wall tension A therapeutic trial of an antimuscarinic drug, given before meals, may be beneficial in patients who have episodes of abdominal pain after eating, but randomized clinical trials demonstrated little, if any, advantage over placebo, and significant risk of mild side effects (Jaiwala et al, 2000) The British Society of Gastroenterology suggests that antidepressants are currently the most effective drugs for IBS In treating underlying depression, antidepressants can modify gut motility and alter visceral nerve responses (Jones et al, 2000) Low doses of

antidepressants were found to be effective in alleviating chronic or severe abdominal pain This effect is independent of their psychotrophic effects and can be achieved at lower doses and a shorter time interval than for that used for patients with psychiatric disorders (Clouse, 1994)

1.5.2.2 Constipation

High fiber diet is frequently recommended for constipation predominant IBS patients Increasing intakeof a range of different dietary "fibers" including those fromcereals, fruits, and vegetables have been shown to increase stool weight and accelerate gut transit (Jones et al, 2000) Wheat bran, at dosesof 10 g to 30 g, is the best known and probably the most effectivefiber supplement Some clinical trials showed that fibers were no better than placebo in overall symptom relief, whereas symptoms such as pain and distension worsened (Snook and Shepherd, 1994) However, most studies reported a significant improvement in constipation if sufficient quantities of fiber are consumed (Cann et al, 1984)

Several new drugs that act on 5HT4 receptors (tegaserod and prucalopride), M3 muscarinic receptors (darifenacine and zamifenacin), and CCK receptors (loxiglumide) appear promising for relief of constipation as well as abdominal pain and bloating in IBS patients (Scarpignato and Pelosini, 1999) These drugs are currently under clinical trials Meanwhile, the current options for treatment of constipation in IBS remain bulk-forming agents (psyllium, methylcellulose), hyperosmotic laxatives (lactulose, sorbitol, and PEG solution), and stool softeners (Ringel et al, 2001)

1.5.2.3 Diarrhea

Loperamide (2-4 mg) is used for treatment of diarrhea-predominant IBS, especially

in patients who also have incontinence As an opioid analogue, loperamide exerts an inhibitory effect on intestinal peristalsis and fluid secretion by interacting with enteric neurons Therefore, it can decrease intestinal transit time, increase intestinal absorption, and strengthen rectal tone (Thompson et al, 2000) It is documented that loperamide significantly ameliorates diarrhea, urgency and faecal soiling (Jailwala et

al, 2000), although it has no effect on other IBS-related symptoms such as pain and bloating (Read et al, 1982)

Chapter 2

Role of Stress in IBS and Gut Axis

Recent years have seen a rapid update in our knowledge of the role of stress in IBS, with the focus now on the brain-gut axis In the following sections, topics about brain-gut axis are discussed Topics include central stress circuitry, hypothalamic-pituitary-adrenal (HPA) axis response, endogenous pain regulation, brain imaging, and changes of bowel functions in motility, and perception

2.2 Effect of stress on brain-gut axis

The GI tract is controlled by a complex network, including the autonomous nervous system (ANS), the CNS, and the enteric nervous system (ENS) that interacts

to establish a bidirectional communication between the brain and the gut, the called “brain-gut axis” These interrelated feedback circuits can influence brain processes and bowel functions affecting pain perception, thoughts and one's appraisal of symptoms, gut perception, secretions, inflammatory responses, and motility Stress is a kind of external or internal stimulus which can activate the brain-gut circuits and induce a series of adaptive and/or maladaptive responses in the CNS and the gut

so-2.2.1 Central circuitry of stress response

Compared with the knowledge on the effects of stress on bowel, the role of central functions in stress and the interactions between neural networks and the gut remain poorly understood Central circuitry of stress response is a neural network composed

of integrative brain structures that generates the response to stress Sawchenko et al pointed out that there are some similar principal circuits underlying the stress response in spite of the different types of stress (Sawchenko et al, 2000) The central stress circuitry may involve the hypothalamus (particular the paraventricular nucleus, PNV), periaqueductal grey, locus coeruleus, amygdale, and limbic forebrain (including the lateral and medial prefrontal cortex) (Monnikes et al, 2001; Sawchenko

et al, 2000) This central network receives input from cortical structures (include medial prefrontal cortex, anterior cingulate cortex, insular etc.) and both visceral and somatic afferents The output of this central stress circuitry is named the emotional

motor system and includes the autonomic nervous system, neuroendocrine (HPA axis), and endogenous pain modulatory system (Mayer, 2000)

A recent study found significant group differences in the frontal brain among healthy controls and IBS patients with respect to the event-related potentials in the brain during exposure to everyday words with emotional content (Blomhoff et al, 2000) Using electroencephalograms (EEG), Nomura et al also showed significantly greater EEG abnormality in the IBS patients (29.2%) than in the controls (4.2%) under mental arithmetic stress and the administration of neostigmine (Nomura et al, 1999) All these data, as well as recent findings generated through brain imaging techniques (see section 2.2.1.4) support the existence of such central stress circuitry

as well as abnormal response of this circuitry in IBS patients under stress situations

2.2.2 Hypothalamic-pituitary-adrenal (HPA) axis response to stress

Activation of the HPA axis is a central component of the stress response This activation is brought about by the release of corticotropin releasing factor (CRF) from the paraventricular nucleus (PVN) of the hypothalamus, which in turn stimulates the release of adrenocorticotropin (ACTH) from the anterior pituitary Circulating ACTH stimulates the release of glucocorticoids (GC) from the adrenal cortex The CRF neurons in the PVN of the hypothalamus control the neuroendocrine response to stress Although the PNV is the primary source of CRF, CRF neurons have also been found in the cortex, limbic and brainstem (Owens and Nemeroff, 1991) The activation of CRF receptors in the brain was shown to activate almost the entire repertoire of behavioral neuroendocrine, autonomic, immunologic, and visceral

responses characteristic of stress in rodents and primates (Habib et al, 2000; Koob et

al, 1999; Webster et al, 1997) Injection of CRF into the CNS of experimental animals produces behavioral and physiological responses similar to those seen in response to acute psychological stress (Mayer, 2000; Tache et al, 1999; Timpl et al, 1998)

Like other neuroendocrine axes, there is also a feedback mechanism in the HPA axis Through CRF receptors located in the medial prefrontal cortex and hippocampus, circulating GC (principal negative feedback mediator) inhibits HPA axis activation and exerts an inhibitory effect on the central stress circuitry (Sternberg et al, 1992) This feedback mechanism observed in response to acute stress can be up or downregulated in various chronic disease states (Mayer, 2000) In certain forms of depression, panic disorder (Gold et al, 1988a) and sexual abuse (DeBellis et al, 1994), hyperactivity of the HPA axis were found in the form of hypercortisolism, which has been considered as the classical form of a generalized stress response escaped from its usual counter-regulation (Gold et al, 1988b) However, there may be a different pattern of stress-induced dysregulation of HPA axis which is characterized by diminishedcortisol levels as a result of a stronger negative feedback inhibition in diarrhea predominant IBS patients In a study, they showed decreased24 hour plasma cortisol, blunted cortisol responses, and normal adrenocorticotropic hormone responses to noxious rectosigmoiddistension (Mayer, 2000) It has been suggested that these patients may have a highly sensitized HPA axis characterizedby decreased basal cortisol levels, increased number of lymphocyte GC receptors, greater suppression of cortisol to dexamethasone, and a more sensitized pituitary gland

(Mayer, 2000) This diminished cortisol response may be associated with increasedcentral CRF responses to stress (Bremner et al, 1997)

2.2.3 Changes in pain modulation

A variety of stressful stimuli has been shown to produce analgesia, a phenomenon often referred to as stress-induced analgesia (SIA) in both humans and experimental animals (Bodnar, 1986; Hayes and Katayama, 1986; Watkins et al, 1982; Basbaum et

al, 1978) Studies using different stressors (e.g cold stress pain, ischemic pain and noxious heat) significantly produced analgesia in healthy subjects (Washington et al, 2000; Willer et al, 1989; Pertovaara et al, 1982) It is suggested that SIA is mediated

by descending pain inhibitory pathways and, dependingon the nature and severity of the stressor, is partially mediated by opioidergic, glutaminergic, and serotonergic systems (Mayer, 2000) The diffuse noxious inhibitory controls (DNIC), also known

as counter-irritation (Willer et al, 1989 and 1999) and the periaqueductal grey rostroventral medulla (RVM) system (Gebhart, 2004) play crucial roles in this endogenous descending pain inhibitory pathways They are central in regulating pain perception and have been shown to be abnormal in IBS associated functional disorders, such as fibromyalgia (Lautenbacher et al, 1997) DNIC relies on spino-bulbo-spinal loops involving ascending pathways in the anterolateral spinal columns, integration in the lower brain stem, and descending influences reaching the dorsal horn neurons

Unlike this “stress inhibiting pain” phenomenon, hypersensitivity to visceral painful distention stress has been reported in patients with IBS (Mayer, 1999)

Recently, a study reported decreased rectal pain thresholds in patients with IBS but not in normal controls during acute laboratorial physical (cold water hand immersion) and psychological (dichotomous listening) stress (Murray et al, 2004) However, the somatic response to stress was found to be different from that of the viscera in IBS Whitehead et al (1990) found that no generalized hypersensitivity to somatic stimulation occurred Furthermore, other studies found IBS patients exhibited cutaneous hypoalgesia during stress (Chang et al, 1999; cook et al, 1987) Such stress induced somatic hypoalgesiaaccompanied by a stress induced visceral hyperalgesia was also found in the rats in response to psychological stressors (Mayer, 2000) One possible explanation for such phenomena is that bothpain facilitatory and inhibitory systems are activated simultaneously in response to stress, the net effect being determined by therelative contribution of these opposing influences (Wei et al, 1999) However, it does not help to explain the finding in healthy controls that physical stress induced visceral hyposensitivity even though it significantly elevated the pain threshold during the stress period (Erckenbrecht et al, 1993)

Taken together, all the above data suggest that stress is associated with modulation

of visceral and somatic sensitivity Furthermore, there is a dysregulation of endogenous pain modulation mechanism in IBS patients compared with normal controls Such alterations in turn could cause visceral hyperalgesia, abdominal symptoms and altered stool habits

2.2.4 Stress and functional brain imaging

Through electroencephalograms (EEG), Nomura et al (1999) showed significantly greater EEG abnormality in the IBS patients (29.2%) than in the controls (4.2%) under mental arithmetic stress Brain imaging techniques such as positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) provide a new and alternative way to “see” what is happening in the brain and measure changes in regional cerebral activity during stress stimulation A study using PET scan to measure vascular perfusion in the brain as an indicator of regional brain activity had indicated that IBS patients may have a different pattern of cerebral response to painful visceral distension stress compared with normal controls (Silverman et al, 1997) The abnormal central responses to visceral painful stimuli were also observed

in many studies using fMRI Some regions such as anterior cingulate cortex, prefrontal cortex (PFC), insular cortex, and thalamus have been observed to respond abnormally (activated or deactivated) during visceral painful distention stress in IBS patients (Mertz et al, 2000; Bernstein et al, 2002; Baciu et al, 1999; Bonaz et al, 2002) Though the findings of the above studies using imaging techniques are not conclusive and are contradictive in some points, they at least proved that there are some abnormalities in central processing to interoceptive stress (visceral pain) However, there is still a lack of research in controlled response to other types of stress such as somatic stress and psychological stress using brain imaging techniques

2.2.5 Effects of stress on gastrointestinal (GI) function

As mentioned before, stress affects brain-gut axis It not only manifests in the changes of central stress circuitry, HPA axis, endogenous pain modulation and brain imaging but also manifests in the GI, the lower part of the brain-gut axis

2.2.5.1 Effects of stress on GI motility

The upper and lower GI tract exhibit differential motor pattern when stimulated by acute stress In common, various acute stressors have been shown to delay gastricemptying (Mayer 2000) while increasing distal colonic motility (Welgan et al, 1988), andaccelerating intestinal transit (Ditto et al, 1998; Martinez et al, 1997) in both healthy humans and animals

Studies in experimental animals showed that various acute stresses (water avoidance, acoustic stimulation, abdominal surgery, operant avoidance) induced delayed gastric emptying (Tache et al, 1993 and 2001) In healthy volunteers, the results are in good agreement with that in animals Various stressors, such as anger, fear, labyrinthine stimulation, painful stimuli, preoperative anxiety, or intense exercise result in an inhibition of gastric motor activity and postprandial emptying (Rao et al, 1998; Tache et al, 1999 and 2001) Antral motility was found to be inhibited in an IBS population Furthermore, studies in patients with functional dyspepsia also showed decreased gastric motility in response to stress (Camilleri et al, 1986) Thus it seems that inhibited gastric motor activity is the principle effect of acute stress in humans and animals

With regard to regional differences, the colon is more responsive to stress than the upper gut (Stam et al, 1995) In rats, stress induced by wrap restraint, cold water swim, or ether slows down intestinal transit (Tache et al, 2001) However, various

tail shock, loud noise, open field test, and restraint at room temperature or in a cold environment) stimulate colonic motility, transit, and/or defecation (Tache et al, 1993 and 2001) Similarly, in healthy human subjects, both physical or psychological stress (hand immersion in cold water, painful stimuli, dichotomous listening test, fear, anxiety, and anger) increases colonic motor activity (Welgan et al, 1988; Rao et al, 1998; Tache et al , 2001) The effects of stress on colonic motility in IBS patients are also in good agreement with that in experiment animals and healthy human subjects Cold pressure, strobe test, mental arithmetic task, fear, and anger all significantly enhance the colonic motor activity (Welgan et al, 1985 and 1988)

Though the above GI motor changes represent the most characteristic patterns induced by various stressors, the underlying mechanism is still not very clear The stress-induced inhibition of gastric emptying and increase in colonic motor activity are suggested to be mediated by CRF acting at the PVN and locus coeruleus and can

be blocked by CRF antagonists and reproduced by CRF administered intraventricularly (Lechner et al, 1997; Gue et al, 1991) Studies showed that cerebral CRF-1 receptors mediated stress induced enhancement of colonic transit, anxiogenic behavioral responses to stress or central administration of CRF (Maillot et al, 2000; Martinez and Tache, 2001; Stickler and Holsboer, 1999), while medullary CRF-2 receptors mediated the stress induced inhibition of gastric emptying (Maillot et al, 2000; Martinez and Tache, 2001) The combined effects of the two CRF receptors and serotonin system may help in the understanding of the different effects of stress

on upper and lower GI tract, as well as the role of CNS in stress modulation Furthermore, many evidences suggested that serotonin (5-HT) is involved in stress

and central CRF mediated changes in GI function, which may cause symptoms in patients with functional GI disorders (Tache et al, 1993; Monnikes et al, 2001) Animal studies showed that stress induced central releasing of CRF was accompanied

by peripherally release of serotonin which stimulated propulsive bowel function through the 5-HT3 receptor (Miyata, 1998; Miyata et al, 1992) However, further studies on the role of other serotonin receptors still need to be carried out to substantiate this finding

2.2.5.2 Effects of stress on GI sensation

Physical and psychological stress can influence gut sensation, which seems to be increased by stress and decreased by relaxation (Drossman et al, 1997) In patients with IBS, alterations in bowel perception were found to be correlated to symptoms over time (Mertz et al, 1995) Because pain is one of the main symptoms in IBS patients, and current published studies on IBS and stress nearly all concentrate in the pain sensation, in the present thesis, we will only discuss on the stress induced changes of pain sensation (see section 2.2.3)

2.2.6 Acute and chronic stress/anxiety animal models

Stress could cause multifacetic debilitating anxiety and depression related disorders (Paterson et al, 2001; Arborelius et al, 1999; McKinney, 1984; Willner, 1997) Since humans and animals share many pathogenic mechanisms in stress response, animal models of stress are useful in elucidating connections between symptoms and biological abnormalities and in suggesting possible treatment

At present, many different acute and chronic stress animal models are available for different research purposes (See table 2-1) Stress-evoked risk assessment is an important domain in anxiety and depression research (Overall, 2000) In animals, these “anxiety-defensive postures” include immobility (wrap restraint), stretched attention (orientation) and “flatback” approach to the stimulus (Kemble & Bolwahnn, 1997; Blanchard et al, 1998; Overall, 2000; 16) There are specific social behaviors in rodents which are very sensitive to anxiety levels (File & Seth, 2003) Homologous to human “social anxiety” state, it is a valuable behavioral model for testing putative anxiotropic manipulations (File & Seth, 2003) Exploratory behaviour (walking, approaches, scanning, etc) is also often studied in the laboratory, in forced (non-escapable) or free (free-choice) situations, as stress-sensitive behavioural parameter to assess anxiety (Cruzio, 2001) Because many forms of human anxiety are based on innate fears of natural dangers, exposure to a predator is known to be a useful model

of “innate” animal anxiety (Newport et al, 2002) However, since the etiologic and pathopsychological mechanisms of stress, anxiety and depression are very complex and still not very clear, current animal models usually been much more simplistic than the disease state they have purported to reflect, and their utility has therefore been limited (Habib et al, 2000)

Among these acute and chronic animal models, wrap restraint stress animal model is one of the commonest animal models used by gastroenterologists in evaluating the effects of stress on bowel function (Smriga et al, 2002; Saito et al, 2002; Million et al, 1999; Williams et al, 1988) Unlike many other stress animal models, it does not result in the formation of ulcers and does not affect gastric

emptying (Williams et al, 1988) There is a strong correlation between stress-induced hormone release and stress-induced intestinal dysfunction over a 24-h period (Williams et al, 1988) Furthermore, there are similarities between the intestinal effects of wrap-restraint stress in rats and intestinal symptoms associated with stress and irritable bowel syndrome in humans Therefore, some gastroenterologists suggested that wrap restraint might be an appropriate animal model in which to study stress-related intestinal dysfunction (Saito et al, 2002; Million et al, 1999; Williams et

al, 1988) Considering of the above factors, this animal model is selected in our study Another reason is because it is quite suitable for our research purpose which aimed to investigate the effects of stress on the intestinal tract However, wrap restraint is only used as an acute stressor Thus, this is a limitation In our study, only the effects of acute stress on bowel motility was investigated Future studies investigating the bowel responses to chronic psychological and physical stressors may be need in animal and human beings to produce a clearer picture of the effects of stress on bowel responses

Table 2-1 Animal models in the study of stress

Holeboard test Inclined or vertical screen test Seed seeking behaviour in hamsters Shock-probe defensive burying Free exploration paradigm

Chronic forced exposure to acute stressors

Learned anxiety (Geller conflict test)

Social

models

Social interaction (File’s) paradigm Maternal or peer separation

Altered group hierarchy Chronic social defeat test

Sensory

models

Exposure to novel or predator odors ZnSO4-induced anosmia

Amputation of vibrissae

Chapter 3

Melatonin and Its Effects on

Brain-Gut Axis and IBS

3.1 Introduction of Melatonin (Synthesis, distribution and metabolism)

3.1.1 Synthesis of melatonin

N-acetyl-5-methoxytryptamine (melatonin) is a derivative of the essential amino acid, tryptophan It was first isolated from the bovine pineal gland and structurally identified by Lerner and colleagues in 1958 (Lerner et al, 1958) In humans and other mammals, the hormone is mainly produced and secreted by the pineal gland according to a pronounced circadian rhythm Extrapinal melatonin production sites such as retina, gastrointestinal tract, urogenital tract do not contribute significantly to blood melatonin levels in mammals, but may be of local importance (Arendt, 1997)

Human pineal gland consists of two types of cells: pinealocytes, which predominate and produce indolamines (mostly melatonin) and peptides, and supporting neuroglial cells The production of melatonin is primarily nocturnal and is controlled by exposure to cycles of light and dark, independent of sleep The synthesis of melatonin is inhibited by exposure to light and stimulated during periods

of darkness by way of a multi-synaptic neural pathway connecting the pineal gland to the external environment via the retina Melatonin levels are low during the day At sunset, the cessation of light triggers neural signals which stimulate the pineal gland

to begin releasing melatonin This rise continues for hours, eventually peaking around

2 a.m (3 a.m for the elderly) after which it steadily declines to minimal levels by morning The delay in timing and decrease in intensity of the melatonin pulse is a manifestation of the aging process (See figure 3-1)