Dopamine transporter imaging in the aged rat. A [ 123 I]FP-CIT SPECT study Aida Ni˜nerola-Baiz´an, Santiago Rojas, N´uria Ro´e-Vellv´e, Francisco Lome˜na, Dom`enec Ros, Javier Pav´ıa PII: DOI: Reference:
S0969-8051(15)00013-X doi: 10.1016/j.nucmedbio.2014.12.020 NMB 7691
To appear in:
Nuclear Medicine and Biology
Received date: Revised date: Accepted date:
1 September 2014 23 December 2014 30 December 2014
Please cite this article as: Ni˜ nerola-Baiz´an Aida, Rojas Santiago, Ro´e-Vellv´e N´ uria, Lome˜ na Francisco, Ros Dom`enec, Pav´ıa Javier, Dopamine transporter imaging in the aged rat. A [123 I]FP-CIT SPECT study, Nuclear Medicine and Biology (2015), doi: 10.1016/j.nucmedbio.2014.12.020
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Title: Dopamine transporter imaging in the aged rat. A [123I]FP-CIT SPECT study.
RI P
T
Abbreviated title: Dopamine transporter imaging in the aged rat
Authors
SC
Aida Niñerola-Baizána,b, Santiago Rojasc,d, Núria Roé-Vellvéb,e, Francisco Lomeñaf,g,h, Domènec
MA NU
Rosa,b,h , Javier Pavíaa,f,h,*
Affiliations: a
Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina
(CIBER-BBN)
Unitat de Biofísica i Bioenginyeria, Facultat de Medicina, Universitat de Barcelona
c
Institut d'Alta Tecnologia, PRBB, CRC Corporació Sanitària
d
Fundació Pasqual Maragall
e
Unidad de Imagen, CIMES, Fundación General de la Universidad de Málaga
f
Servei de Medicina Nuclear, Hospital Clínic de Barcelona
g
Centro de Investigación Biomédica en Red de Salud Mental (CIBERSAM)
h
Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS)
AC
CE
PT
ED
b
Corresponding author: Javier Pavía Servei de Medicina Nuclear - Hospital Clínic C/ de Villarroel 170 - 08036 Barcelona, Spain +34932275400 - [email protected]
1
ACCEPTED MANUSCRIPT Abstract Introduction: Rodent models are extensively used to assess the biochemical and physiological
T
changes associated with aging. They play a major role in the development of therapies for age-
RI P
related pathologies such as Parkinson's disease. To validate the usefulness of these animal models in aging or age-related disease research, the consistency of cerebral aging processes across species must
SC
be evaluated. The dopaminergic system seems particularly susceptible to the aging process. One of the results of this susceptibility is a decline in striatal dopamine transporter (DAT) availability.
MA NU
Methods: We sought to ascertain whether similar age changes could be detected in-vivo in rats, using molecular imaging techniques such as single photon emission computed tomography (SPECT) with [123I]FP-CIT.
ED
Results: A significant decrease of 17.21% in the striatal specific uptake ratio was observed in the aged rats with respect to the young control group.
PT
Conclusions: Our findings suggest that age-related degeneration in the nigrostriatal track is similar in humans and rats, which supports the use of this animal in models to evaluate the effect of aging on
CE
the dopaminergic system.
AC
Advances in Knowledge and Implications for patient Care: Our findings indicate that age-related degeneration in the nigrostriatal track is similar in humans and rats and that these changes can be monitored in vivo using small animal SPECT with [123I]FP-CIT, which could facilitate the translational research in rat models of age related disorders of dopaminergic system.
Keywords Aging, Dopamine, Dopamine Transporter (DAT), [123I]FP-CIT, rat, Single Positron Emission Computed Tomography (SPECT).
2
ACCEPTED MANUSCRIPT Abbreviations Dopamine
DAT
Dopamine Transporter
PD
Parkinson's Disease
PET
Positron Emission Tomography
ROI
Region Of Interest
SNpc
Substantia Nigra pars compacta
SPECT
Single Photon Emission Computed Tomography
SUR
Specific Uptake Ratio
AC
CE
PT
ED
MA NU
SC
RI P
T
DA
3
ACCEPTED MANUSCRIPT 1. INTRODUCTION Motor function declines during aging in animals and humans [1,2]. This motor dysfunction is
T
associated with the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc),
RI P
resulting in low levels of dopamine (DA) in the human basal ganglia [3] and changes in dopamine transporter (DAT) activity and dopamine receptor density [4]. To date, both postsynaptic and
SC
presynaptic elements of the nigrostriatal dopaminergic system have been probed for age-related
MA NU
changes.
A number of studies have evaluated the effect of aging in this neural system using molecular imaging techniques such as single photon emission computed tomography (SPECT) and positron emission
ED
tomography (PET). Some of them were focused on postsynaptic receptors (D1 and D2) localized in the striatum and cerebral cortex, others evaluated pre-synaptic targets in the nigral neurons. While
PT
results on the D1-like DA receptor remain inconclusive, molecular imaging studies point to a substantial loss of D2-like receptors with age, reporting age-dependent losses around a 4.6-8%
CE
decrease per decade at the level of striatum [5,6]. Unlike postsynaptic receptors, presynaptic markers
AC
can provide direct information about the nigrostriatal cells. In particular DAT, that is responsible for reuptake of DA from synaptic cleft, is associated with presynaptic dopamine function integrity of dopaminergic nigrostriatal neuron. The study of DAT ligand binding is of great interest, as it has proved to be a sensitive indicator of disturbance in nigrostriatal function. To date, several PET or SPECT studies have reported aging-associated decreases in DAT [7-12] and its findings have been confirmed in humans by post-mortem evaluation of the nigrostriatal track [3].
Rodent models are used extensively in the assessment of biochemical and physiological process associated with aging as well as for the evaluation of therapies for age-related pathological conditions. However, to serve as a valid animal model for research into brain aging processes, 4
ACCEPTED MANUSCRIPT interspecies differences in neurotransmission systems must be explored. Research shows that rodents experience age-associated changes in some neurotransmission systems that are completely
T
comparable with those found in humans. For example, similar reductions in the binding of [11C]-
RI P
raclopride in the striatum has been found in aged rats and humans [5, 6, 13]. However, the evolution of other neurotransmission systems does not always follow the same pattern or even the same
SC
direction. One example of this are PET ligands for the serotonin transporter that increase its binding
MA NU
in aged rats but, surprisingly, decrease in humans [14-15].
Using molecular imaging techniques to map the effects of age on the different neurotransmission systems in rodents could pave the way for longitudinal studies to assess the effect of
ED
pharmacological and behavioral interventions in normal and pathological brain aging. In this way, understanding the aging process of the rodent brain could be essential to translate to humans our
PT
findings from animal models of neurodegenerative disorders.
CE
DAT imaging with [123I]FP-CIT SPECT has been used in experimental models of Parkinson's
AC
disease (PD) [16-17], and it is widely used for the study of the integrity of the nigrostriatal system in the diagnosis of PD and related pathologies [18]. Recently, its use to discriminate dementia with Lewy bodies from Alzheimer’s disease has also been validated [18-19]. Quantitative declines in DAT binding have been documented, reporting a 5.5% to 8% decrease in striatal receptor binding potential per decade in healthy humans using PET and SPECT techniques [7, 9-12]. To our knowledge, the effect of aging on nigrostriatal system has not been yet evaluated in rodents with small animal SPECT using the DAT ligand [123I]FP-CIT.
In the present study, our aim was to quantify the density of DAT in young and old rats, using in vivo SPECT imaging with the radiotracer [123I]FP-CIT. With this approach, we want to explore first if the 5
ACCEPTED MANUSCRIPT changes in the DAT density in rats are similar than those described in humans. And second, validate
T
the use of SPECT with the radiotracer [123I]FP-CIT to quantify in vivo the DAT density in aged rats.
RI P
2. MATERIAL AND METHODS 2.1. Animals
SC
Twelve male Oncins France Strain A (OFA) rats (Charles River, Barcelona, Spain) were used in this study, six young adult animals and six aged animals [20]. At the time of SPECT acquisition, the
MA NU
younger animals weighed 263.8 ± 34.2g (mean±SD) and the aged ones 630.8 ± 77.4g. At that time, the control group was 9.5 ± 0.3 weeks old and the older group was 74.4 ± 0.3 weeks old. All animals were housed on a 12h light/dark cycle with free access to food and water until the SPECT was
ED
carried out. All animal experiments were performed in accordance with the National Institute of Health guide for care and use of laboratory animals and were approved by local government
2.2. SPECT imaging
PT
authorities.
CE
All scans were performed with a dedicated small-animal SPECT system [21] (2mm intrinsic
AC
resolution) equipped with a low energy high resolution parallel-hole collimator. A dose of approximately 90 MBq of [123I]FP-CIT (specific activity 314.5 ± 5.2 MBq/mmol; radiochemical purity of 98.5 ± 0.1%) was injected intravenously into the tail vein. Animals were imaged at 30 min post injection, once this reversible tracer reached the pseudoequilibrium state [22]. For dose injection and SPECT acquisition, animals were maintained under anaesthesia with isoflurane at 1.5% vaporized in oxygen. Data were acquired for 60 min in a step-and-shoot mode over a circular orbit in angular steps of 6° (60 projections, 60 s/projection) and a rotation radius of 30 mm. 2.3. Image quantification and statistical analysis
6
ACCEPTED MANUSCRIPT Projections were filtered using a 2D-Butterworth filter (cut-off frequency 3.13 cm-1, order 5) and reconstructed using a filtered back projection algorithm with a ramp filter with a 128×128×100
T
matrix size and a 0.32×0.32×0.32 mm3 voxel size.
RI P
Correction for attenuation and scatter was not applied. These degrading effects are only relevant in case of significant between-group differences in the size of the scanned organs. In rat brain, these
SC
differences between young and aged animals were only apparent in the antero-posterior axis (average 1 mm longer brains in the aged group) and were below 0.25 mm in the rest of the directions. Such
MA NU
dimensions are completely irrelevant in terms of the attenuation of gamma rays in biological tissues. After reconstruction, SPECT images were visually coregistered to an anatomical rat brain atlas [23] using a rigid body transformation to verify the anatomical location of the striatal uptake. Afterwards,
ED
striatum was delineated in the SPECT images and a reference region was selected in the cerebellum
PT
because of the absence of DAT in this brain structure.
Striatal binding was evaluated by calculating the Specific Uptake Ratio (SUR), defined as SUR = (S-
CE
B)/B, where S is the concentration activity in the striatum and B is that of the background reference
AC
region with non-specific uptake. This is the preferred quantification method when a reference region is available in the brain and is the most suitable method when there are differences in body mass, injected dose or physiologic parameters between the groups. Differences of weight that could affect the amount of radiotracer delivered into the brain, do not affect this parameter because weight has no effect on either the specific affinity in striatum or non-specific affinity. This makes specific uptake ratio the parameter of choice in [123I]FP-CIT SPECT in experimental and clinical studies. For those readers familiar with PET technology, where the most common parameter to be evaluated is the Standard Uptake Value (SUV), it should be pointed out that the SUR parameter supplies the same information as the ratio between SUV in the striatum (SUVS) and SUV in the background
7
ACCEPTED MANUSCRIPT reference region (SUVB). In fact, SUR =SUVS/SUVB-1, as can be derived directly by considering the two definitions.
T
SUR values were compared across groups by means of Student’s t-tests. P-values below P
S0969-8051(15)00013-X doi: 10.1016/j.nucmedbio.2014.12.020 NMB 7691
To appear in:
Nuclear Medicine and Biology
Received date: Revised date: Accepted date:
1 September 2014 23 December 2014 30 December 2014
Please cite this article as: Ni˜ nerola-Baiz´an Aida, Rojas Santiago, Ro´e-Vellv´e N´ uria, Lome˜ na Francisco, Ros Dom`enec, Pav´ıa Javier, Dopamine transporter imaging in the aged rat. A [123 I]FP-CIT SPECT study, Nuclear Medicine and Biology (2015), doi: 10.1016/j.nucmedbio.2014.12.020
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Title: Dopamine transporter imaging in the aged rat. A [123I]FP-CIT SPECT study.
RI P
T
Abbreviated title: Dopamine transporter imaging in the aged rat
Authors
SC
Aida Niñerola-Baizána,b, Santiago Rojasc,d, Núria Roé-Vellvéb,e, Francisco Lomeñaf,g,h, Domènec
MA NU
Rosa,b,h , Javier Pavíaa,f,h,*
Affiliations: a
Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina
(CIBER-BBN)
Unitat de Biofísica i Bioenginyeria, Facultat de Medicina, Universitat de Barcelona
c
Institut d'Alta Tecnologia, PRBB, CRC Corporació Sanitària
d
Fundació Pasqual Maragall
e
Unidad de Imagen, CIMES, Fundación General de la Universidad de Málaga
f
Servei de Medicina Nuclear, Hospital Clínic de Barcelona
g
Centro de Investigación Biomédica en Red de Salud Mental (CIBERSAM)
h
Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS)
AC
CE
PT
ED
b
Corresponding author: Javier Pavía Servei de Medicina Nuclear - Hospital Clínic C/ de Villarroel 170 - 08036 Barcelona, Spain +34932275400 - [email protected]
1
ACCEPTED MANUSCRIPT Abstract Introduction: Rodent models are extensively used to assess the biochemical and physiological
T
changes associated with aging. They play a major role in the development of therapies for age-
RI P
related pathologies such as Parkinson's disease. To validate the usefulness of these animal models in aging or age-related disease research, the consistency of cerebral aging processes across species must
SC
be evaluated. The dopaminergic system seems particularly susceptible to the aging process. One of the results of this susceptibility is a decline in striatal dopamine transporter (DAT) availability.
MA NU
Methods: We sought to ascertain whether similar age changes could be detected in-vivo in rats, using molecular imaging techniques such as single photon emission computed tomography (SPECT) with [123I]FP-CIT.
ED
Results: A significant decrease of 17.21% in the striatal specific uptake ratio was observed in the aged rats with respect to the young control group.
PT
Conclusions: Our findings suggest that age-related degeneration in the nigrostriatal track is similar in humans and rats, which supports the use of this animal in models to evaluate the effect of aging on
CE
the dopaminergic system.
AC
Advances in Knowledge and Implications for patient Care: Our findings indicate that age-related degeneration in the nigrostriatal track is similar in humans and rats and that these changes can be monitored in vivo using small animal SPECT with [123I]FP-CIT, which could facilitate the translational research in rat models of age related disorders of dopaminergic system.
Keywords Aging, Dopamine, Dopamine Transporter (DAT), [123I]FP-CIT, rat, Single Positron Emission Computed Tomography (SPECT).
2
ACCEPTED MANUSCRIPT Abbreviations Dopamine
DAT
Dopamine Transporter
PD
Parkinson's Disease
PET
Positron Emission Tomography
ROI
Region Of Interest
SNpc
Substantia Nigra pars compacta
SPECT
Single Photon Emission Computed Tomography
SUR
Specific Uptake Ratio
AC
CE
PT
ED
MA NU
SC
RI P
T
DA
3
ACCEPTED MANUSCRIPT 1. INTRODUCTION Motor function declines during aging in animals and humans [1,2]. This motor dysfunction is
T
associated with the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc),
RI P
resulting in low levels of dopamine (DA) in the human basal ganglia [3] and changes in dopamine transporter (DAT) activity and dopamine receptor density [4]. To date, both postsynaptic and
SC
presynaptic elements of the nigrostriatal dopaminergic system have been probed for age-related
MA NU
changes.
A number of studies have evaluated the effect of aging in this neural system using molecular imaging techniques such as single photon emission computed tomography (SPECT) and positron emission
ED
tomography (PET). Some of them were focused on postsynaptic receptors (D1 and D2) localized in the striatum and cerebral cortex, others evaluated pre-synaptic targets in the nigral neurons. While
PT
results on the D1-like DA receptor remain inconclusive, molecular imaging studies point to a substantial loss of D2-like receptors with age, reporting age-dependent losses around a 4.6-8%
CE
decrease per decade at the level of striatum [5,6]. Unlike postsynaptic receptors, presynaptic markers
AC
can provide direct information about the nigrostriatal cells. In particular DAT, that is responsible for reuptake of DA from synaptic cleft, is associated with presynaptic dopamine function integrity of dopaminergic nigrostriatal neuron. The study of DAT ligand binding is of great interest, as it has proved to be a sensitive indicator of disturbance in nigrostriatal function. To date, several PET or SPECT studies have reported aging-associated decreases in DAT [7-12] and its findings have been confirmed in humans by post-mortem evaluation of the nigrostriatal track [3].
Rodent models are used extensively in the assessment of biochemical and physiological process associated with aging as well as for the evaluation of therapies for age-related pathological conditions. However, to serve as a valid animal model for research into brain aging processes, 4
ACCEPTED MANUSCRIPT interspecies differences in neurotransmission systems must be explored. Research shows that rodents experience age-associated changes in some neurotransmission systems that are completely
T
comparable with those found in humans. For example, similar reductions in the binding of [11C]-
RI P
raclopride in the striatum has been found in aged rats and humans [5, 6, 13]. However, the evolution of other neurotransmission systems does not always follow the same pattern or even the same
SC
direction. One example of this are PET ligands for the serotonin transporter that increase its binding
MA NU
in aged rats but, surprisingly, decrease in humans [14-15].
Using molecular imaging techniques to map the effects of age on the different neurotransmission systems in rodents could pave the way for longitudinal studies to assess the effect of
ED
pharmacological and behavioral interventions in normal and pathological brain aging. In this way, understanding the aging process of the rodent brain could be essential to translate to humans our
PT
findings from animal models of neurodegenerative disorders.
CE
DAT imaging with [123I]FP-CIT SPECT has been used in experimental models of Parkinson's
AC
disease (PD) [16-17], and it is widely used for the study of the integrity of the nigrostriatal system in the diagnosis of PD and related pathologies [18]. Recently, its use to discriminate dementia with Lewy bodies from Alzheimer’s disease has also been validated [18-19]. Quantitative declines in DAT binding have been documented, reporting a 5.5% to 8% decrease in striatal receptor binding potential per decade in healthy humans using PET and SPECT techniques [7, 9-12]. To our knowledge, the effect of aging on nigrostriatal system has not been yet evaluated in rodents with small animal SPECT using the DAT ligand [123I]FP-CIT.
In the present study, our aim was to quantify the density of DAT in young and old rats, using in vivo SPECT imaging with the radiotracer [123I]FP-CIT. With this approach, we want to explore first if the 5
ACCEPTED MANUSCRIPT changes in the DAT density in rats are similar than those described in humans. And second, validate
T
the use of SPECT with the radiotracer [123I]FP-CIT to quantify in vivo the DAT density in aged rats.
RI P
2. MATERIAL AND METHODS 2.1. Animals
SC
Twelve male Oncins France Strain A (OFA) rats (Charles River, Barcelona, Spain) were used in this study, six young adult animals and six aged animals [20]. At the time of SPECT acquisition, the
MA NU
younger animals weighed 263.8 ± 34.2g (mean±SD) and the aged ones 630.8 ± 77.4g. At that time, the control group was 9.5 ± 0.3 weeks old and the older group was 74.4 ± 0.3 weeks old. All animals were housed on a 12h light/dark cycle with free access to food and water until the SPECT was
ED
carried out. All animal experiments were performed in accordance with the National Institute of Health guide for care and use of laboratory animals and were approved by local government
2.2. SPECT imaging
PT
authorities.
CE
All scans were performed with a dedicated small-animal SPECT system [21] (2mm intrinsic
AC
resolution) equipped with a low energy high resolution parallel-hole collimator. A dose of approximately 90 MBq of [123I]FP-CIT (specific activity 314.5 ± 5.2 MBq/mmol; radiochemical purity of 98.5 ± 0.1%) was injected intravenously into the tail vein. Animals were imaged at 30 min post injection, once this reversible tracer reached the pseudoequilibrium state [22]. For dose injection and SPECT acquisition, animals were maintained under anaesthesia with isoflurane at 1.5% vaporized in oxygen. Data were acquired for 60 min in a step-and-shoot mode over a circular orbit in angular steps of 6° (60 projections, 60 s/projection) and a rotation radius of 30 mm. 2.3. Image quantification and statistical analysis
6
ACCEPTED MANUSCRIPT Projections were filtered using a 2D-Butterworth filter (cut-off frequency 3.13 cm-1, order 5) and reconstructed using a filtered back projection algorithm with a ramp filter with a 128×128×100
T
matrix size and a 0.32×0.32×0.32 mm3 voxel size.
RI P
Correction for attenuation and scatter was not applied. These degrading effects are only relevant in case of significant between-group differences in the size of the scanned organs. In rat brain, these
SC
differences between young and aged animals were only apparent in the antero-posterior axis (average 1 mm longer brains in the aged group) and were below 0.25 mm in the rest of the directions. Such
MA NU
dimensions are completely irrelevant in terms of the attenuation of gamma rays in biological tissues. After reconstruction, SPECT images were visually coregistered to an anatomical rat brain atlas [23] using a rigid body transformation to verify the anatomical location of the striatal uptake. Afterwards,
ED
striatum was delineated in the SPECT images and a reference region was selected in the cerebellum
PT
because of the absence of DAT in this brain structure.
Striatal binding was evaluated by calculating the Specific Uptake Ratio (SUR), defined as SUR = (S-
CE
B)/B, where S is the concentration activity in the striatum and B is that of the background reference
AC
region with non-specific uptake. This is the preferred quantification method when a reference region is available in the brain and is the most suitable method when there are differences in body mass, injected dose or physiologic parameters between the groups. Differences of weight that could affect the amount of radiotracer delivered into the brain, do not affect this parameter because weight has no effect on either the specific affinity in striatum or non-specific affinity. This makes specific uptake ratio the parameter of choice in [123I]FP-CIT SPECT in experimental and clinical studies. For those readers familiar with PET technology, where the most common parameter to be evaluated is the Standard Uptake Value (SUV), it should be pointed out that the SUR parameter supplies the same information as the ratio between SUV in the striatum (SUVS) and SUV in the background
7
ACCEPTED MANUSCRIPT reference region (SUVB). In fact, SUR =SUVS/SUVB-1, as can be derived directly by considering the two definitions.
T
SUR values were compared across groups by means of Student’s t-tests. P-values below P
