- Email: [email protected]
Ligands for peripheral benzodiazepine binding sites in glial cells
Brain Research Reviews 48 (2005) 207 – 210 www.elsevier.com/locate/brainresrev
Review
Ligands for peripheral benzodiazepine binding sites in glial cells Michael Kassioua,b,c,*, Steven R. Meiklea,d, Richard B. Banatia,d a
Ramaciotti Centre for Brain Imaging, Brain and Mind Research Institute (BMRI), University of Sydney, NSW 2006, Australia b Department of PET and Nuclear Medicine, Royal Prince Alfred Hospital, NSW 2050, Australia c Department of Pharmacology, University of Sydney, NSW 2006, Australia d School of Medical Radiation Sciences, University of Sydney, NSW 1825, Australia Accepted 9 December 2004 Available online 22 January 2005
Abstract Within the diseased brain, glial cells and in particular, microglia, express a multimeric protein complex termed bperipheral benzodiazepine binding sites (PBBS)Q or bperipheral benzodiazepine receptor (PBR)Q. The expression of the PBBS is dependent on the functional state of the cell and in glial cells is triggered by a wide range of activating stimuli. In the healthy brain, the PBBS are nearly absent with the notable exception of the choroid plexus, ependymal layer, perivascular cells, central canal, possibly certain nuclei in the brainstem and layers in the cerebellum where a constitutive presence of the PBBS is found. Likewise, areas that due to the absence of the blood–brain barrier contain more active glial cells, such as the pituitary gland, or the area postrema at floor of the 4th ventricle show a degree of constitutive expression. The tight correlation of the parenchymal de novo expression of the PBBS with the presence of activated glial cells, that in turn are usually only found in tissue affected by progressive disease, establishes the PBBS as a generic marker for the detection and measurement of active disease processes in the brain. Specific radioligands of the PBBS for use in positron emission tomography (PET) may thus provide a sensitive in vivo index of neuropathological activity. Whilst prototype ligands for the PBBS are available, future research needs to focus on the development of new ligands with improved pharmacodynamic properties and the ability to discriminate between the different, still insufficiently understood functional states of the peripheral benzodiazepine receptor complex. D 2004 Elsevier B.V. All rights reserved. Theme: Disorders of the nervous system Topic: Degenerative disease Keywords: Peripheral benzodiazepine receptor; Activated microglia; Neurodegenerative disease
Contents 1. Introduction. . . . . . . . . . . . . . . . . 2. The peripheral benzodiazepine binding sites 3. Molecular structure and function . . . . . . 4. PBBS in neurodegenerative disease . . . . References . . . . . . . . . . . . . . . . . . . .
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* Corresponding author. Department of PET and Nuclear Medicine, Royal Prince Alfred Hospital, Missenden Road Camperdown, NSW 2050, Australia. Fax: +61 2 9515 6381. E-mail address: [email protected] (M. Kassiou). 0165-0173/$ - see front matter D 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.brainresrev.2004.12.010
208
M. Kassiou et al. / Brain Research Reviews 48 (2005) 207–210
1. Introduction One index useful in delineating areas of active disease often well before any other overt pathological or obvious structural changes is the presence of activated microglia, the brain’s indigenous population of tissue macrophages that is ubiquitously distributed throughout the central nervous system. In normal brain paenchyma, microglia are in a seemingly bdown-regulatedQ state, whereas during disease or injury they rapidly respond to the pathological event, upregulating or expressing de novo a wide array of molecules, including the PBBS [12]. The diagnostic value of detecting activated microglia for the accurate spatial localization of active disease processes, acute or chronic, is also a reflection of the fact that unlike astrocytes, microglia are isolated cells that do not respond like a syncytium within which an activation process can spread diffusely beyond the original site of activation. Therefore, in cases of neuronal injury, microglial activation tends to remain strictly localized to the injured neuronal axon and its retrograde and anterograde projection areas. Thus, the bvisualizationQ of activated microglia is particularly suited as a target for non-invasive in vivo monitoring of disease progression in the brain using positron emission tomography (PET) [1].
2. The peripheral benzodiazepine binding sites (PBBS) The peripheral benzodiazepine binding sites (PBBS) is anatomically and pharmacologically distinct from the wellknown central benzodiazepine receptor (CBR) found in the CNS. The two binding sites differ in their anatomical distribution, pharmacological and biochemical functions, and relative affinities for specific ligands. The CBRs are the site at which classical benzodiazepine ligands, such as diazepam, exert their anxiolytic, anticonvulsant, and muscle relaxant effects through the g-aminobutyric acid (GABAA) complex. The PBBS on the other hand are involved in regulatory processes and metabolic functions pertaining to the tissue in which they are present. They are abundantly expressed in peripheral organs, such as the kidney, heart, and the steroid hormone producing cells of the adrenal cortex, testis, and the ovaries. In the brain, unlike the CBR counterparts, the PBBS are found only in non-neuronal tissue of the ependyma, epithelial cells of the choroid plexus and glial cells, which have been activated by a pathological stimulus. Within the cell, the PBBS are primarily localized in mitochondria where their distribution is closely correlated with the mitochondrial enzymes cytochrome oxidase, succinate dehydrogenase, and monoamine oxidase [19]. Within mitochondria, the PBBS are located in the outer mitochondrial membrane. Ligand binding studies have revealed a second subcellular location of PBBS [7] in the liver, heart, adrenals, and testes that appears to be associated with a cell
surface membrane fraction. Furthermore, the fact that PBBS are also found in red blood cells which lack mitochondria [13] indicates that the PBBS are also localized in non-mitochondrial fractions. For microglia, this issue has not yet been specifically addressed. Although the benzodiazepine diazepam exhibits high affinity binding to both the PBBS and CBR binding sites, the two can be distinguished through the use of other selective ligands. Ligands, such as flumazenil, iomazenil, and clonazepam, bind to the CBR with nanomolar affinity and with less than micromolar affinity for the PBBS [16]. Conversely, ligands such as Ro 5-4864 (7-chloro-1,3dihydro-1-methyl-5(4V-chlorophenyl)-2H-1,4-benzodiazepine-2-one) and PK11195 (1-(2-chlorophenyl)-N-methyl-N(1-methyl-propyl)-3-isoquinoline carboxamide), which exhibit no obvious anxiolytic activity, bind to the PBBS with nanomolar affinity but have at least a 1000-fold lower affinity for the CBR [17].
3. Molecular structure and function The PBBS are multimeric protein complexes composed of an 18-kDa subunit, a 32-kDa subunit that functions as a voltage-dependent anion channel (VDAC) or porin and a 30-kDa subunit that functions as an adenine nucleotide carrier (ANC) [9]. Isoquinoline ligands such as PK 11195 that bind to the PBBS interact specifically with the 18-kDa subunit [15], whereas PBBS-specific ligands based on the benzodiazepine structure (e.g., Ro 5-4864) bind to a site consisting of both VDAC and the 18-kDa isoquinoline subunit. The ANC appears to bind PBBS-type specific benzodiazepines. The exact arrangement of these protein subunits to form the PBBS complex is still not known. In studies carried out on MA-10 Leydig cell mitochondria, it was shown that the 18-kDa PBBS protein is organized in clusters of four to six molecules in association with one VDAC molecule leading to the formation of bsingle poresQ [15] also called the Mitochondrial Permeability Transition pore. More recently, it was suggested that the PBBS are associated with other proteins and that the solubilized receptor is actually part of a bigger 200 kDa complex [8]. Photoaffinity labeling with PK 14105 (analogue of PK 11195) has led to the identification of an another 10-kDa protein associated with the 18-kDa units of the PBBS [6] through a new protein, called PRAX-1 (for peripheral benzodiazepine receptor associated protein 1) which is involved in linking and dimerizing these subunits. Therefore, it appears that different molecular structures may interact with different binding domains on this receptor complex. Unlike the CBR, the functions and roles of the PBBS are far less understood. Furthermore, the activity spectrum of the PBBS is very wide. Hence, an important criterion for elucidating the functions of the PBBS must take particular note of their tissue and subcellular localization. Some of
M. Kassiou et al. / Brain Research Reviews 48 (2005) 207–210
these functions include calcium homeostasis, cholesterol transport and steroidogenesis, heme and porphyrin transport, cell proliferation, cellular respiration, and cellular immunity [10]. One of the most significant findings is the regulation of the steroid hormone production induced by pituitary trophic hormones. The key step in this process is the conversion of cholesterol into pregnelone by cytochrome P-450 side-chain cleavage (P-450scc) enzyme, which is located on the inner mitochondrial membrane. The transport of cholesterol from the outer to the inner mitochondrial membrane is rate limiting in steroid hormone synthesis. PBBS ligands not only stimulate steroid synthesis in adrenal, placental, testicular, and ovarian cells, but also in the brain and a strong correlation between potency of stimulation and affinity of ligands for the receptor has been reported [14,15]. Ro5-4864, for example, enhances cholesterol transport from cell cytoplasm to inner mitochondrial membrane. In the brain, it appears to be mainly neurosteroids, such as pregnelone, that might indirectly modulate synaptic (GABA-ergic) transmission.
4. PBBS in neurodegenerative disease Perhaps one of the most significant findings is the increased PBBS density in a variety of neurodegenerative conditions and other forms of neuronal injuries. Increased PBBS density has been observed postmortem in the brains of patients suffering from Parkinson’s [11] and Alzheimer’s disease (AD) [7] using [3H]Ro5-4864. In the latter study, a two- to threefold increase in density was observed in Broca’s area, central and pre-central gyri while changes were found to be insignificant in other regions, including the temporal gyri where the greatest neuronal loss is expected. Furthermore, increases in PBBS density appeared not to correlate with the activities of the marker enzyme choline acetyl transferase (CAT) and acetylcholinesterase [11]. However, both of these studies involved the use of [3H]Ro5-4864, a tracer shown earlier to have a low affinity for human PBBS. Subsequently, in studies using [3H]PK11195 instead of [3H]Ro5-4864, a 120% increase in PBBS density was observed in temporal cortex and a moderate increase in the frontal cortex (reflecting an increase in receptor density rather than a reduced [3H]Ro5-4864 affinity), when compared to normal control patients. In these studies, decreased CAT was observed in both regions [4]. Since the PBBS in the CNS occur mainly in glial cells, it was suggested that the elevated levels in PBBS density reflect brain damage and gliosis. Studies of tissue from 14 patients with adult onset Huntington’s disease showed a significant increase (51%) in the binding of [3H]Ro5-4864 in the putamen but not to caudate and globus pallidus compared to 18 controls [18]. These findings were similar to the pathology in rat brains with selective neuronal death in the striatum following intrastriatal injection of kainic acid.
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Significant changes in PBBS density were also observed in postmortem brain tissue from other neurological and neuropsychiatric conditions. In postmortem studies of cerebrovascular disease using [3H]PK11195, a sevenfold increase in PBBS density was seen in the periphery of the infarcted areas 7 and 22 days after the stroke [5]. Although there was no correlation between PBBS density and the age of the lesion, the increased PBBS density corresponded well with the perimeter of the lesion as confirmed by histology. Similarly, studies on postmortem material from three patients with multiple sclerosis, showed a tenfold increase in PBBS density around the periphery of the plaques compared to that in normal white matter with the extent of demyelination correlating well with increased PBBS density [6]. In schizophrenic patients under anti-dopaminergic treatment, a reduction in PBBS density in platelets has been reported [20]. Here, the significance of the observation remained unclear due to the complexities of dopamine–PBBS interactions. The above discrepancies are potentially due to significant differences in the various binding domains making up the PBBS complex. This in particular highlights the need for further ligand refinement in affinity, selectivity, and structure in order to determine which components of this receptor complex are associated with PBBS function in disease states. These ligands are also needed to address the issue of cellular selectivity in greater detail and the fact that–at least in microglia–the PBBS may provide a good indicator of the transition from a brestingQ into an bactivatedQ state, but does not reflect a further important cellular event, namely, the transformation into phagocytic macrophages [3]. Current ligands, such as PK11195, do not adequately reflect the severity of disease once the cellular activation has occurred. Thus, maps of PK11195 binding do not sufficiently discriminate between areas in which glial PBBS expression is associated with tissue destruction and subsequent atrophy from those in which the glial response is part of potentially adaptive or restorative processes [1,2]. References [1] R.B. Banati, Visualising microglia activation in vivo, Glia 40 (2002) 206 – 217. [2] R.B. Banati, Brain plasticity and microglia: is transsynaptic glial activation in the thalamus after limb denervation linked to cortical plasticity and central sensitisation, J. Physiol. (Paris) 96 (2002) 289 – 299. [3] R.B. Banati, R. Myers, G.W. Kreutzberg, PK (dperipheral benzodiazepineT)-binding sites in the CNS indicate early and discrete brain lesions: microautoradiographic detection of [3H]PK11195 binding to activated microglia, J. Neurocytol. 26 (1997) 77 – 82. [4] J. Benavides, D. Fage, C. Carter, B. Scatton, Peripheral type benzodiazepine binding sites are a sensitive indirect index of neuronal damage, Brain Res. 421 (1987) 167 – 172. [5] J. Benavides, P. Cornu, T. Dennis, A. Dubois, J.J. Hauw, E.T. MacKenzie, V. Sazdovitch, B. Scatton, Imaging of human brain lesions with an omega 3 site radioligand, Ann. Neurol. 24 (1988) 708 – 712.
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[6] J. Blahos, M.E. Whalin, K.E. Krueger, Identification and purification of a 10-kilodalton protein associated with mitochondrial benzodiazepine receptors, J. Biol. Chem. 270 (1995) 20285 – 20291. [7] A. Doble, J. Benavides, O. Ferris, P. Bertrand, J. Menager, N. Vaucher, M.C. Burgevin, A. Uzan, C. Gueremy, G. Le Fur, Dihydropyridine and peripheral type benzodiazepine binding sites: subcellular distribution and molecular size determination, Eur. J. Pharmacol. 119 (1985) 153 – 167. [8] S. Galiegue, O. Jbilo, T. Combes, E. Bribes, P. Carayon, G. Le Fur, P. Casellas, Cloning and characterization of PRAX-1. A new protein that specifically interacts with the peripheral benzodiazepine receptor, J. Biol. Chem. 274 (1999) 2938 – 2952. [9] M. Garnier, A.B. Dimchev, N. Boujrad, J.M. Price, N.A. Musto, V. Papadopoulos, In vitro reconstitution of a functional peripheral-type benzodiazepine receptor from mouse Leydig tumor cells, Mol. Pharmacol. 45 (1994) 201 – 211. [10] J.D. Hirsch, C.F. Beyer, L. Malkowitz, C.C. Loullis, A.J. Blume, Characterization of ligand binding to mitochondrial benzodiazepine receptors, Mol. Pharmacol. 35 (1989) 164 – 172. [11] E.G. McGeer, E.A. Singh, P.L. McGeer, Peripheral-type benzodiazepine binding in Alzheimer disease, Alzheimer Dis. Assoc. Disord. 2 (1988) 331 – 336. [12] L.B. Moran, D.C. Duke, F.E. Turkheimer, R.B. Banati, M.B. Graeber, Towards a transcriptome definition of microglial cells, Neurogenetics 5 (2004) 95 – 108. [13] F. Owen, M. Poulter, J.L. Waddington, R.D. Masha, T.J. Crow, [3H]Ro5-4864 and flunitrazepam binding in kainate-lesioned rat
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[19] [20]
striatum and in temporal cortex of brains from patients with senile dementia of the Alzheimer type, Brain Res. 278 (1983) 373 – 375. V. Papadopoulos, A.G. Mukhin, E. Costa, K.E. Krueger, The peripheral-type benzodiazepine receptor is functionally linked to Leydig cell steroidogenesis, J. Biol. Chem. 265 (1990) 3772 – 3779. V. Papadopoulos, H. Amri, N. Boujrad, C. Cascio, M. Culty, M. Garnier, M. Hardwick, H. Li, B. Vidic, A.S. Brown, J.L. Reversa, J.M. Bernassau, K. Drieu, Peripheral benzodiazepine receptor in cholesterol transport and steroidogenesis, Steroids 62 (1997) 21 – 28. V.W. Pike, C. Halldin, C. Crouzel, L. Barre, D.J. Nutt, S. Osman, F. Shah, D.R. Turton, S.L. Waters, Radioligands for PET studies of central benzodiazepine receptors and PK (peripheral benzodiazepine) binding sites—current status, Nucl. Med. Biol. 20 (1993) 503 – 525. S. Selleri, F. Bruni, C. Costagli, A. Costanzo, G. Guerrini, G. Ciciani, B. Costa, C. Martini, 2-Arylpyrazolo[1,5-a]pyrimidin-3-yl acetamides. New potent and selective peripheral benzodiazepine receptor ligands, Bioorg. Med. Chem. 9 (2001) 2661 – 2671. H. Schoemaker, M. Morelli, P. Deshmukh, H.I. Yamamura, [3H]Ro5-4864 benzodiazepine binding in the kainate lesioned striatum and Huntington’s diseased basal ganglia, Brain Res. 248 (1982) 396 – 401. A. Verma, S.H. Snyder, Peripheral type benzodiazepine receptors, Annu. Rev. Pharmacol. Toxicol. 29 (1989) 307 – 322. R. Weizman, Z. Tanne, L. Karp, S. Tyano, M. Gavish, Peripheral-type benzodiazepine-binding sites in platelets of schizophrenics with and without tardive dyskinesia, Life Sci. 39 (1986) 549 – 555.
Review
Ligands for peripheral benzodiazepine binding sites in glial cells Michael Kassioua,b,c,*, Steven R. Meiklea,d, Richard B. Banatia,d a
Ramaciotti Centre for Brain Imaging, Brain and Mind Research Institute (BMRI), University of Sydney, NSW 2006, Australia b Department of PET and Nuclear Medicine, Royal Prince Alfred Hospital, NSW 2050, Australia c Department of Pharmacology, University of Sydney, NSW 2006, Australia d School of Medical Radiation Sciences, University of Sydney, NSW 1825, Australia Accepted 9 December 2004 Available online 22 January 2005
Abstract Within the diseased brain, glial cells and in particular, microglia, express a multimeric protein complex termed bperipheral benzodiazepine binding sites (PBBS)Q or bperipheral benzodiazepine receptor (PBR)Q. The expression of the PBBS is dependent on the functional state of the cell and in glial cells is triggered by a wide range of activating stimuli. In the healthy brain, the PBBS are nearly absent with the notable exception of the choroid plexus, ependymal layer, perivascular cells, central canal, possibly certain nuclei in the brainstem and layers in the cerebellum where a constitutive presence of the PBBS is found. Likewise, areas that due to the absence of the blood–brain barrier contain more active glial cells, such as the pituitary gland, or the area postrema at floor of the 4th ventricle show a degree of constitutive expression. The tight correlation of the parenchymal de novo expression of the PBBS with the presence of activated glial cells, that in turn are usually only found in tissue affected by progressive disease, establishes the PBBS as a generic marker for the detection and measurement of active disease processes in the brain. Specific radioligands of the PBBS for use in positron emission tomography (PET) may thus provide a sensitive in vivo index of neuropathological activity. Whilst prototype ligands for the PBBS are available, future research needs to focus on the development of new ligands with improved pharmacodynamic properties and the ability to discriminate between the different, still insufficiently understood functional states of the peripheral benzodiazepine receptor complex. D 2004 Elsevier B.V. All rights reserved. Theme: Disorders of the nervous system Topic: Degenerative disease Keywords: Peripheral benzodiazepine receptor; Activated microglia; Neurodegenerative disease
Contents 1. Introduction. . . . . . . . . . . . . . . . . 2. The peripheral benzodiazepine binding sites 3. Molecular structure and function . . . . . . 4. PBBS in neurodegenerative disease . . . . References . . . . . . . . . . . . . . . . . . . .
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* Corresponding author. Department of PET and Nuclear Medicine, Royal Prince Alfred Hospital, Missenden Road Camperdown, NSW 2050, Australia. Fax: +61 2 9515 6381. E-mail address: [email protected] (M. Kassiou). 0165-0173/$ - see front matter D 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.brainresrev.2004.12.010
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M. Kassiou et al. / Brain Research Reviews 48 (2005) 207–210
1. Introduction One index useful in delineating areas of active disease often well before any other overt pathological or obvious structural changes is the presence of activated microglia, the brain’s indigenous population of tissue macrophages that is ubiquitously distributed throughout the central nervous system. In normal brain paenchyma, microglia are in a seemingly bdown-regulatedQ state, whereas during disease or injury they rapidly respond to the pathological event, upregulating or expressing de novo a wide array of molecules, including the PBBS [12]. The diagnostic value of detecting activated microglia for the accurate spatial localization of active disease processes, acute or chronic, is also a reflection of the fact that unlike astrocytes, microglia are isolated cells that do not respond like a syncytium within which an activation process can spread diffusely beyond the original site of activation. Therefore, in cases of neuronal injury, microglial activation tends to remain strictly localized to the injured neuronal axon and its retrograde and anterograde projection areas. Thus, the bvisualizationQ of activated microglia is particularly suited as a target for non-invasive in vivo monitoring of disease progression in the brain using positron emission tomography (PET) [1].
2. The peripheral benzodiazepine binding sites (PBBS) The peripheral benzodiazepine binding sites (PBBS) is anatomically and pharmacologically distinct from the wellknown central benzodiazepine receptor (CBR) found in the CNS. The two binding sites differ in their anatomical distribution, pharmacological and biochemical functions, and relative affinities for specific ligands. The CBRs are the site at which classical benzodiazepine ligands, such as diazepam, exert their anxiolytic, anticonvulsant, and muscle relaxant effects through the g-aminobutyric acid (GABAA) complex. The PBBS on the other hand are involved in regulatory processes and metabolic functions pertaining to the tissue in which they are present. They are abundantly expressed in peripheral organs, such as the kidney, heart, and the steroid hormone producing cells of the adrenal cortex, testis, and the ovaries. In the brain, unlike the CBR counterparts, the PBBS are found only in non-neuronal tissue of the ependyma, epithelial cells of the choroid plexus and glial cells, which have been activated by a pathological stimulus. Within the cell, the PBBS are primarily localized in mitochondria where their distribution is closely correlated with the mitochondrial enzymes cytochrome oxidase, succinate dehydrogenase, and monoamine oxidase [19]. Within mitochondria, the PBBS are located in the outer mitochondrial membrane. Ligand binding studies have revealed a second subcellular location of PBBS [7] in the liver, heart, adrenals, and testes that appears to be associated with a cell
surface membrane fraction. Furthermore, the fact that PBBS are also found in red blood cells which lack mitochondria [13] indicates that the PBBS are also localized in non-mitochondrial fractions. For microglia, this issue has not yet been specifically addressed. Although the benzodiazepine diazepam exhibits high affinity binding to both the PBBS and CBR binding sites, the two can be distinguished through the use of other selective ligands. Ligands, such as flumazenil, iomazenil, and clonazepam, bind to the CBR with nanomolar affinity and with less than micromolar affinity for the PBBS [16]. Conversely, ligands such as Ro 5-4864 (7-chloro-1,3dihydro-1-methyl-5(4V-chlorophenyl)-2H-1,4-benzodiazepine-2-one) and PK11195 (1-(2-chlorophenyl)-N-methyl-N(1-methyl-propyl)-3-isoquinoline carboxamide), which exhibit no obvious anxiolytic activity, bind to the PBBS with nanomolar affinity but have at least a 1000-fold lower affinity for the CBR [17].
3. Molecular structure and function The PBBS are multimeric protein complexes composed of an 18-kDa subunit, a 32-kDa subunit that functions as a voltage-dependent anion channel (VDAC) or porin and a 30-kDa subunit that functions as an adenine nucleotide carrier (ANC) [9]. Isoquinoline ligands such as PK 11195 that bind to the PBBS interact specifically with the 18-kDa subunit [15], whereas PBBS-specific ligands based on the benzodiazepine structure (e.g., Ro 5-4864) bind to a site consisting of both VDAC and the 18-kDa isoquinoline subunit. The ANC appears to bind PBBS-type specific benzodiazepines. The exact arrangement of these protein subunits to form the PBBS complex is still not known. In studies carried out on MA-10 Leydig cell mitochondria, it was shown that the 18-kDa PBBS protein is organized in clusters of four to six molecules in association with one VDAC molecule leading to the formation of bsingle poresQ [15] also called the Mitochondrial Permeability Transition pore. More recently, it was suggested that the PBBS are associated with other proteins and that the solubilized receptor is actually part of a bigger 200 kDa complex [8]. Photoaffinity labeling with PK 14105 (analogue of PK 11195) has led to the identification of an another 10-kDa protein associated with the 18-kDa units of the PBBS [6] through a new protein, called PRAX-1 (for peripheral benzodiazepine receptor associated protein 1) which is involved in linking and dimerizing these subunits. Therefore, it appears that different molecular structures may interact with different binding domains on this receptor complex. Unlike the CBR, the functions and roles of the PBBS are far less understood. Furthermore, the activity spectrum of the PBBS is very wide. Hence, an important criterion for elucidating the functions of the PBBS must take particular note of their tissue and subcellular localization. Some of
M. Kassiou et al. / Brain Research Reviews 48 (2005) 207–210
these functions include calcium homeostasis, cholesterol transport and steroidogenesis, heme and porphyrin transport, cell proliferation, cellular respiration, and cellular immunity [10]. One of the most significant findings is the regulation of the steroid hormone production induced by pituitary trophic hormones. The key step in this process is the conversion of cholesterol into pregnelone by cytochrome P-450 side-chain cleavage (P-450scc) enzyme, which is located on the inner mitochondrial membrane. The transport of cholesterol from the outer to the inner mitochondrial membrane is rate limiting in steroid hormone synthesis. PBBS ligands not only stimulate steroid synthesis in adrenal, placental, testicular, and ovarian cells, but also in the brain and a strong correlation between potency of stimulation and affinity of ligands for the receptor has been reported [14,15]. Ro5-4864, for example, enhances cholesterol transport from cell cytoplasm to inner mitochondrial membrane. In the brain, it appears to be mainly neurosteroids, such as pregnelone, that might indirectly modulate synaptic (GABA-ergic) transmission.
4. PBBS in neurodegenerative disease Perhaps one of the most significant findings is the increased PBBS density in a variety of neurodegenerative conditions and other forms of neuronal injuries. Increased PBBS density has been observed postmortem in the brains of patients suffering from Parkinson’s [11] and Alzheimer’s disease (AD) [7] using [3H]Ro5-4864. In the latter study, a two- to threefold increase in density was observed in Broca’s area, central and pre-central gyri while changes were found to be insignificant in other regions, including the temporal gyri where the greatest neuronal loss is expected. Furthermore, increases in PBBS density appeared not to correlate with the activities of the marker enzyme choline acetyl transferase (CAT) and acetylcholinesterase [11]. However, both of these studies involved the use of [3H]Ro5-4864, a tracer shown earlier to have a low affinity for human PBBS. Subsequently, in studies using [3H]PK11195 instead of [3H]Ro5-4864, a 120% increase in PBBS density was observed in temporal cortex and a moderate increase in the frontal cortex (reflecting an increase in receptor density rather than a reduced [3H]Ro5-4864 affinity), when compared to normal control patients. In these studies, decreased CAT was observed in both regions [4]. Since the PBBS in the CNS occur mainly in glial cells, it was suggested that the elevated levels in PBBS density reflect brain damage and gliosis. Studies of tissue from 14 patients with adult onset Huntington’s disease showed a significant increase (51%) in the binding of [3H]Ro5-4864 in the putamen but not to caudate and globus pallidus compared to 18 controls [18]. These findings were similar to the pathology in rat brains with selective neuronal death in the striatum following intrastriatal injection of kainic acid.
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Significant changes in PBBS density were also observed in postmortem brain tissue from other neurological and neuropsychiatric conditions. In postmortem studies of cerebrovascular disease using [3H]PK11195, a sevenfold increase in PBBS density was seen in the periphery of the infarcted areas 7 and 22 days after the stroke [5]. Although there was no correlation between PBBS density and the age of the lesion, the increased PBBS density corresponded well with the perimeter of the lesion as confirmed by histology. Similarly, studies on postmortem material from three patients with multiple sclerosis, showed a tenfold increase in PBBS density around the periphery of the plaques compared to that in normal white matter with the extent of demyelination correlating well with increased PBBS density [6]. In schizophrenic patients under anti-dopaminergic treatment, a reduction in PBBS density in platelets has been reported [20]. Here, the significance of the observation remained unclear due to the complexities of dopamine–PBBS interactions. The above discrepancies are potentially due to significant differences in the various binding domains making up the PBBS complex. This in particular highlights the need for further ligand refinement in affinity, selectivity, and structure in order to determine which components of this receptor complex are associated with PBBS function in disease states. These ligands are also needed to address the issue of cellular selectivity in greater detail and the fact that–at least in microglia–the PBBS may provide a good indicator of the transition from a brestingQ into an bactivatedQ state, but does not reflect a further important cellular event, namely, the transformation into phagocytic macrophages [3]. Current ligands, such as PK11195, do not adequately reflect the severity of disease once the cellular activation has occurred. Thus, maps of PK11195 binding do not sufficiently discriminate between areas in which glial PBBS expression is associated with tissue destruction and subsequent atrophy from those in which the glial response is part of potentially adaptive or restorative processes [1,2]. References [1] R.B. Banati, Visualising microglia activation in vivo, Glia 40 (2002) 206 – 217. [2] R.B. Banati, Brain plasticity and microglia: is transsynaptic glial activation in the thalamus after limb denervation linked to cortical plasticity and central sensitisation, J. Physiol. (Paris) 96 (2002) 289 – 299. [3] R.B. Banati, R. Myers, G.W. Kreutzberg, PK (dperipheral benzodiazepineT)-binding sites in the CNS indicate early and discrete brain lesions: microautoradiographic detection of [3H]PK11195 binding to activated microglia, J. Neurocytol. 26 (1997) 77 – 82. [4] J. Benavides, D. Fage, C. Carter, B. Scatton, Peripheral type benzodiazepine binding sites are a sensitive indirect index of neuronal damage, Brain Res. 421 (1987) 167 – 172. [5] J. Benavides, P. Cornu, T. Dennis, A. Dubois, J.J. Hauw, E.T. MacKenzie, V. Sazdovitch, B. Scatton, Imaging of human brain lesions with an omega 3 site radioligand, Ann. Neurol. 24 (1988) 708 – 712.
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