Journal of Chronic Fatigue Syndrome, Vol. 4(3) 1998 Relationship Between SPECT Scans and Buspirone Tests in Patients with ME/CFS John Richardson, MB, BS Durval Campos Costa, MD, MSc, PhD ABSTRACT. The purpose of this exercise was to study the relationship between the detail shown on the SPECT brain scans with those seen in the buspirone tests. Thirty-nine patients are included in this study. These patients were selected from a large number who had been referred to Dr. Richardson from various parts of the country by their doctors because of a tentative diagnosis of ME/CFS. All the selected patients were confirmed by Dr. Richardson as suffering from ME/CFS taking into account the subjective scoring methods, clinical examination, virology and buspirone tests. This study is an attempt to link together the results of the previously described techniques to investigate possible areas of impaired cellular function in brain which may have purely neuroneural effects or possibly neurohormonal effects. All patients within this study displayed hypoperfusion in some brain area as shown by their SPECT scans (see Appendix, Table 1.1). * Thirty-five (90%) showed hypoperfusion in the regions comprising: * Twenty-four (62%) in the Brain Stem * Twenty (51%) in the Caudate Nuclei * Nine (23%) showed hypoperfusion in both Brain Stem and Caudate Nuclei regions * Thirty (77%) cases demonstrated hypoperfusion in the regions comprising: * Twenty-four (62%) in the Temporal Lobes * Twelve (31%) in the Parietal Lobes * Nine (23%) in the Frontal Lobes The significance of these results is to confirm that there is actual evidence of neurological dysfunction which results in the continuing morbidity in these ME/CFS patients. The completion of this buspirone test and SPECT scan can be deemed to be basic complementary evidence for the positive diagnosis of ME/CFS. _______________ John Richardson is in private practice, England. Durval Campos Costa is affiliated with the Institute of Nuclear Medicine, The Middlesex Hospital, London. Address correspondence to: Dr. John Richardson, "Belle Vue," Grange Road, Ryton, Tyne & Wear, NE40 3LU, England. _______________ INTRODUCTION Non-invasive techniques, such as SPECT brain scans were used by Durval Campos Costa et al. (1) to investigate the possible areas of brain affected in myalgic encephalomyelitis/chronic fatigue syndrome(ME/CFS). On the other hand, biochemical measurements of neurohormonal, such as the Post Buspirone Prolactin Cortisol variation test (2) were conducted by Dr. Richardson following the initial work of Bakheit et al. (3). It appeared reasonable to look at the relationship between these two tests and speculate on the results and what they may suggest in terms of related biological function, and then to perform both tests on a group of patients and study the outcome relationship. Initial considerations suggested that in the SPECT scans, the fields of hypoperfusion which were shown to be significant demonstrated localised areas with diminished metabolic requirements. This could be related to diminished cell function which occurred as a result of mitochondrial dysfunction, possibly due to viral infection. This has been well demonstrated by Behan and coworkers. It should be noted that mitochondria, as the name implies, are the "nuclear power house" of the cell, from "mitosis" to "apoptosis." During cell life, this mitochondrial "power" results in the generation of energy in the form of ion gradients and ATP synthesis. Mitochondria mediate energy generation through the oxidation of food material which has passed through the blood-brain-barrier (BBB). To this end, mitochondria also contain the enzymes for the Krebs and other fatty acid cycles, and thus govern the respiratory cell pathway of oxygen. Therefore, mitochondrial dysfunction results in a decrease in cell respiration. For instance, abnormal carnitine metabolism in ME/CFS patients is one of the energy metabolic dysfunctions in mitochondria. Carnitine itself is an amino acid used for the transport of fatty acids across the cell membrane to the matrix, a process which is catalysed by the co-enzyme carnitine palmitoxyl transferase. The resulting decrease in oxidation is no doubt the reason for the lack of perfusion requirements demonstrated in the SPECT scans. Mitochondria contain DNA and RNA by means of which they replicate. This relates to the development of cells with the formation of deutoplasm and a large increase in protoplasm and mitochondria, which leads to the formation and future function of specialised cells. Other physiological aspects that may underlie the SPECT abnormalities seen in ME/CFS should be considered, such as the anatomical divisions of the brain with enormous variations in function. The latter variations are seen in both the neuroneural and neurohormonal control of the whole body. Consideration should be given to the BBB, as this possibly has some effect on the means of access to the higher centres of viruses or toxins as well as normal nourishment, with the areas above the BBB being possibly more protected than those beneath. However, viruses and other material, e.g., toxins and certain drugs which are fat soluble, may pass this BBB. The microglial cells are of mesodermal origin whilst the astrocytes and oligodendrocytes, like the rest of the brain, are of ectodermal origin. These microglial cells can migrate and, like the general reticular-endothelial system, they function as the autoimmune system above the BBB. This autoimmune function when activated, will exercise itself in relation to any infected cellular material in the CNS and this will result in a response related to the cellular substance. Microglial cells disperse through the cerebrospinal fluid and this dispersion is probably the reason for the expansion of the Virchov-Robin spaces, which, in turn, may be the reason for the "cuffing" seen around the retinal veins in some ME/CFS cases. The latter cuffing has been postulated to be the reason for the so called unidentified bright objects (UBOs) seen on MRI scans. We have seen this and Royce Biddle, MD, of Stockton, California has alluded to this in his chapter in the book, by the Nightingale Research Foundation (4). The cellular pathological response that results from the activities described above may be postulated to underlie the reduced metabolic demands of cells in the loci affected, and may become evident as hypoperfusion on SPECT scans. Therefore, the various areas adversely affected might be delineated by the hypoperfusion shown on a SPECT scan. In the current study, we have focused on the limbic system, which extends from the hippocampal formation of the temporal lobes, the fornix and mammillary bodies, to the anterior nucleus of the thalamus, cingula, septal area and the orbital surface of the frontal lobes. The limbic system comprises the hippocampus and dentate gyroseptal areas and amygdala, which are associated with olfaction but are of greater importance in other autonomic functions such as emotion and behaviour. The hippocampus is 5 cm long and is an elevation of the floor of the inferior horn of the lateral ventricle. Its efferent pathway is the fornix with efferent olfactory fibres to the mammillary bodies. The fibres from the hippocampus form the alveus which converge to form the fimbria. Traced backwards the fimbria form the posterior column of the fornix beneath the corpus callosum. These columns are connected by transverse fibres-the hippocampal commissure. The fornix is an efferent pathway and its fibres, relayed to the mammillary bodies, pass to the anterior nucleus of the thalamus by the mammillothalamic tract, and to the brain stem by the mammillotegmental tract. The latter network shows the intricacies of the association areas in the region defined as the "brain stem." There is a close relationship between the hypothalamic-mammillary area and the brain stem reticular formation. The limbic system and the reticular formation are closely involved in relation to memory function which is often impaired in ME/CFS. Viral infection or trauma resulting in encephalitis in these areas may have long-term effects. In considering the information obtained from SPECT scans, the areas of hypoperfusion can thus be considered in relation to the function associated with that brain area. The hypoperfusion is a result rather than a cause of the abnormalities seen in ME/CFS. The outcome of the SPECT scans and buspirone tests is also considered in relation to the clinical signs and symptoms presented at examination (see Appendix, Tables 1, 2 and 3). METHODS Patient Selection Thirty-nine patients are included in this study, seventeen males and twenty-two females with ages ranging from 14 to 58 years. These patients were selected from a large number who had been referred to Dr. Richardson from various parts of United Kingdom by their doctors because of a tentative diagnosis of ME/CFS. All the selected patients were confirmed by Dr. Richardson as suffering from ME/CFS, taking into account the subjective scoring methods, clinical examination, and recent, positive, virology and buspirone tests. The subjective scoring methods included the Newcastle Research Group (NRG) Score Chart, the CDC and Oxford criteria. The NRG Score Chart, as shown in Table 4 (see Appendix), has been used for over four decades and differentiates between central CNS and peripheral muscle fatigue as well as cognitive disorders. A score of 13 indicates a positive indication of ME/CFS. The Hamilton Score Chart test was used to exclude any patients with psychiatric illness from this study. Full clinical examination and serological testing was performed to exclude, as far as possible, any other systemic disease as a cause of the illness. In all the cases within this study, evidence obtained by antibody testing was positive for previous enterovirus infection. In most cases, the VP1 or PCR test for enterovirus was undertaken and was positive, usually on multiple occasions over many months. The patients in this group had all been ill for years and one or two, such as F22, for more than a decade. F22 is an example of a few who, if their SPECT scans had been performed during the darker days of their illness might well have demonstrated a more severe degree of hypoperfusion in the areas affected. In the case of F22, the symptoms relating to the ME/ CFS have now diminished and her recovery is estimated to be about 80%. As shown, her SPECT scan only reveals diminished hypoperfusion to the left temporal area which is the area relating to the epilepsy. It is significant that the amelioration of the purely neuroneural symptoms seen in epilepsy which indicate nerve cell pathology are less likely to resolve by cell regeneration and this is well demonstrated in poliomyelitis. Thus, it could be appropriate to repeat SPECT scans after apparent recovery in other cases. In the present series, not all SPECT scans were done at the height of the illness and, therefore, it is a matter of conjecture as to what the results may have been prior to some degree of recovery. Buspirone Stimulation Studies It has been suggested that one of the major effects of persistent virus infections in the production of disorders such as myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is on the hypothalamus (3). Buspirone, which is one of the anxiolytic drugs of the azopyrone group, causes a release of prolactin by stimulation of serotonin 5-hydroxytryptophan (5-HT) receptors. The buspirone-prolactin response was studied in a subgroup of patients with ME/CFS and evidence of persistent enteroviral infection, as shown by the repeated detection of the group-specific protein of enteroviruses, VP], in the blood. Family controls who were asymptomatic were studied at the same time. In addition to the response to buspirone, diurnal variations in cortisol and prolactin levels were studied. It was found that the patients with ME/CFS had much greater rises in prolactin levels one hour after buspirone compared to controls. Cortisol levels were elevated in the patients, but the rise was not significantly different between the two groups. There was a significant association between the pattern of sleep disturbance, which we speak of as the OWL syndrome, and the rise in pre- and post-buspirone prolactin levels. The buspirone tests were performed by Dr. Richardson according to the protocol published earlier (2). The blood samples were subsequently analysed by Mr. Dennis Peel in the Department of Biochemistry, Queen Elizabeth Hospital, Gateshead, Tyne and Wear. SPECT Scans Brain perfusion studies were carried out by D. C. Costa at the Institute of Nuclear Medicine, University College London Medical School, London, UK. Each patient was given Tc-99m-HMPAO (Ceretec(@) intravenously and data acquisition followed 15-30 minutes later using a brain dedicated triple head gamma camera (GE Neurocam). Data acquisition, processing and analysis followed previously described routine protocols. A written report and graph plot of the quantified regional brain perfusion expressed as radioactivity ratios was issued for each patient. The quantification was always based on the comparison against a database of normal volunteers matched for age comprising 40 studies of 26 males and 14 females with age ranging from 21 to 59 years of age. The graph plot (Figure 1) displays the mean perfusion ratios for each brain region using the cerebellum as denominator, as well as the 96% confidence limits of the population, not of the mean ( + 2SD to - 2SD). All brain regions presenting ratios below 2SD from the mean were considered hypoperfused. RESULTS The purpose of this exercise was to study the relationship between the detail shown on the SPECT brain scans with those seen in the buspirone tests. The brain can be subdivided into areas which vary in function but are closely integrated. This exercise was aimed at looking into the functions of brain loci which may have their ongoing effects mediated by neurohormonal activity (via the pituitary or other glands), and other areas which have their far reaching effects mediated by neuroneural activity (via outgoing neurological pathways), and to see if these brain regions can be defined and possibly related to the clinical illness. FIGURE 1. An example of the graphical output from the SPECT scan. [Figure 1 can be viewed at: http://www.co-cure.org/private/brain/Figure1.htm ] Analysis of SPECT scans from ME/CFS patients reveals the following characteristics: * Thirty-nine (100%) showed hypoperfusion in some brain area * Thirty-five (90%) showed hypoperfusion: * Twenty-four (62%) in the Brain Stem, and * Twenty (51%) in the Caudate Nuclei * Nine (23%) showed hypoperfusion in both Brain Stem and Caudate Nuclei regions * Thirty (77%) cases demonstrated hypoperfusion: * Twenty-four (62%) in the Temporal Lobes, * Twelve (31%) in the Parietal Lobes, and * Nine (23%) in the Frontal Lobes Buspirone Test In the earlier paper (2), the following observation was made with regard to the rise in prolactin level one hour after administration of the buspirone, "A rise of 250% and upwards encompasses 26 out of 30 patients and a rise of less than this defines a similar proportion of controls, assuring a reliability for the distinction of 87%." Visual Cortex In none of the patients tested was hypoperfusion evident in the visual cortical areas (RVC and LVC). The visual cortex includes the greater part of the occipital lobe on both sides and is subdivided into visuosensory and visuopsychic areas. Visuocortical areas are in the walls of the post calcarine sulcus and extend on to the surface of the cuneus above and lingual gyrus below. There are four areas of cells with varying functions. The outer and inner granular layers receive fibres from the optic radiation and therefore become the screen upon which vision is projected. The ganglionic layer sends fibres to the corpus quadrigeminum and adjoining area of the midbrain. The cells in these layers are profuse, small and closely packed, and number a tenth part of the whole of the cerebral cortex. Colour, size, form, motion and other modalities of visuosensory recognition occur here. Recognition and identification of objects requires the operation of the visuopsychic area which surrounds the visuosensory area. Thus, this area correlates visual impressions with past experience and orientates objects in spatial relations. The fact that the visuocortical areas, which act as the receptor screen for the optic radiation, appear to be spared in SPECT scans may suggest that the visual difficulties which ME cases have, could be secondary to some aberration in the function of the areas which receive the fibres from the visual cortex, such as the superior quadrigeminal body which is concerned with reflex movements of the head and eyes in response to visual stimuli. Obviously, the association pathways in the brain which function on a neuroneural or neurohormonal basis and the complex interactions of their physiological activity is beyond the scope of this paper but should be seriously considered as a matter for ongoing research. Epilepsy Three cases of epilepsy are included within this limited study, others have been seen who are not included but have also succumbed following the viral infection. Reading-induced and sonagenic epilepsy are recorded. EEGs in two of these patients have shown slow wave activity in the right temporal lobes in addition to the usual epileptogenic focus in the left temporal lobes. Similar results have also been identified in work by Richard L. Bruno, PhD, of the Kessler Institute for Rehabilitation in New Jersey (6). DISCUSSION Clinical Examination Over the last four decades, the diagnosis for ME/CFS was based upon the classical criteria of history and clinical examination and these still remain the cornerstone and should be conducted with great care. The written histories which were presented to Dr. Richardson by patients described these symptoms and formed the basis for the NRG Score Chart. This also showed the persistence of illness long after the initial illness commenced. The information gathered from the Score Chart delineated pathology as being of central CNS or more peripheral regions. As shown in Table 2, Physical Stress in this series was shown to apply to 5 male patients but to no females and chiefly related to physical habits and work commitments. Three of these males have, sometime in their illness, shown signs of myocarditis. This confirms previous research which showed the myocardial effects of physical stress in patients with viral illness. Viral Investigation It was apparent that many of these cases which occurred in both epidemic and endemic clusters, followed a viral infection. Serological testing was routinely performed and repeated over a number of occasions. This gave evidence in many cases for persisting viral infection. These investigations did not, however, delineate the resulting pathology following the viral infection and, unhappily, there was clinical evidence of persisting illness. Thus, it became mandatory to develop investigations which would enable us to define the pathological abnormalities in these areas. These investigations included cardiac, peripheral muscle, liver, renal and other tissue biopsies. In all of these, evidence of persistent viral infection was found but this is beyond the remit of this paper. Pathological Investigation There was clinical evidence of neuroneural- and neurohormonal-mediated pathology, and initial investigations into these areas owe much to the work of D. C. Costa, A. M. O. Bakheit, P. O. Behan and G. T. Dinan. For this reason, both cortisol and prolactin studies were performed which related to their normal diurnal variation and response to buspirone (2). This study demonstrated no significant change in normal diurnal variation of prolactin or cortisol but a marked rise in prolactin secretion one hour after the administration of buspirone. It was found that patients with ME/ CFS had much greater rises in prolactin levels one hour after buspirone as compared to controls. In the latter group, there was either an insignificant rise or occasionally a slight fall, whereas a mane to post buspirone rise of 250% and upwards was seen in the ME/CFS patients. All the patients considered within this study demonstrated such a rise except for case F22 (see Table 3). F22 was known to suffer from epilepsy and, therefore, was given a reduced 20 mg dose of buspirone to avoid unnecessary side effects. It is reasonable to expect that the resulting 81% rise would have been greater than 250% had the full 50 mg dose been administered. SPECT Scans The present work is an attempt to link together the results of the previously described techniques to investigate possible areas of impaired cellular function in brain which may have purely neuroneural effects or possibly neurohormonal effects. The neuroneural effects possibly could be related to upper cortical regions such as the frontal, temporal and parietal regions. The neurohormonal effects were presumed to indicate changes such as hypothalamic-pituitary-adrenal axis abnormalities. These have been shown to result in adverse changes relating to the stability of bodily states such as temperature, blood pressure, heart rate and even thalamic pain and emotion thresholds. The brain stem, as well as the hypothalamic areas, is also involved in relation to these effects. All patients within this study displayed hypoperfusion in some brain area as shown by their SPECT scans (see Table 1.1). As is shown in Table 1.2, there is a marked difference between the profiles of male and female patients within this trial regarding the percentages showing hypoperfusion in various brain regions. Male patients showed a significantly higher incidence of hypoperfusion in the temporal and caudate areas, while in females the frontal and parietal areas were more commonly involved. There was no significant difference in the brainstem area. The relevance of this bias is not apparent and would need to be confirmed in a larger sample of cases before warranting further analysis. Conclusions The significance of the SPECT scan and buspirone test results reported herein is to confirm that there is actual evidence of neurological dysfunction which results in the continuing morbidity in ME/CFS patients. It should not be assumed that these are the only neuroneural or neurohormonal abnormalities present in ME/CFS. Other areas, such as the neuroneural optic radiation (as previously mentioned) or the neurohormonal thymus gland, may also be involved, a possibility which warrants further investigation. Thus, the apparent sparing of the visual cortex on SPECT scans does not preclude any abnormality of the ongoing optic radiation to the corpus quadrigeminum which is possibly the reason for the focusing difficulties in this group of patients. The completion of the buspirone test and SPECT scan can be deemed to be basic complementary evidence for the positive diagnosis of ME/CFS. The current implications of the findings reported for treatment are that we should engender a more sympathetic attitude to the ME/CFS patient, and should pursue this in relation to possible reparative treatment which may involve immunological and other mechanisms outside the remit of this paper. However, in neurological conditions, healing and repair are notoriously imperfect. REFERENCES 1. Costa D. C. et al. Brainstem perfusion is impaired in chronic fatigue syndrome. Q J Med 1995; 88:767-773. 2. Richardson J. Disturbance of hypothalamic function and evidence for persistent enteroviral infection in patients with chronic fatigue syndrome. J Chronic Fatigue Syndrome 1995; 1(2):59-66. 3. Bakheit A. M. O., Behan P. O., Dinan G. T., Gray C. E., O'Keene V. G. Upregulation of 5-HT receptors in patients with PVFS. Brit Med J 1992; 304:110-112. 4. Biddle R. The Clinical and Scientific basis of ME/CFS, Nightingale Research Foundation, Chapter 48, 1992. APPENDIX TABLE 1.1. Results of Trial Patients SPECT Scans and Buspirone Tests [This table can be read at: http://www.co-cure.org/private/brain/Table1-1.htm ] TABLE 1.2. Variation Between Male and Female Patients Regarding the Areas Showing Hypoperfusion [This table can be read at: http://www.co-cure.org/private/brain/Table1-2.htm ] TABLE 2. Clinical Signs of the Patients [This table can be read at: http://www.co-cure.org/private/brain/Table2.htm ] TABLE 3. Results of Trial Patients Prolactin/Buspirone Tests [This table can be read at: http://www.co-cure.org/private/brain/Table3.htm ] TABLE 4. Newcastle Research Group ME/CFS Score Chart [This table can be read at: http://www.co-cure.org/private/brain/Table4.htm ]