Validation of a Tracer Kinetic Model for the Quantification of 5-HT 2A Receptors in Human Brain with [ 11 C]MDL 100,907

Subjects and Medication

Five normal volunteers (four males aged 37, 55, 62 and 63 years and one female of 59 years) gave written informed consent to be included in the study. They were screened by an experienced psychiatrist (ZB) to exclude any past or current history of psychiatric illness using the Structured Clinical Interview for DSM-IV disorders (SCID; First et al, 1997). They were also screened using the Hamilton Depression Rating Scale (HDRS; Hamilton 1960) (mean score 0) and the Beck Depression Inventory (BDI; Beck et al, 1961) (mean score 0). Subjects were included only if they had not been on any medication for at least 6 months before the scan, did not abuse alcohol or illicit drugs and did not have any significant current or history of medical or neurologic illness.

Each subject had two PET scans using [¹¹C]MDL 100,907 at the same time of the day with an interval of at least 1 week between the two scans. Subjects were given a single tablet of mirtazapine 30 mg approximately 90 mins before the start of the acquisition of PET data on one of the two occasions in a randomized open label design. In healthy volunteers, mirtazapine is rapidly absorbed after a single dose and its peak plasma concentration is reached within 1 to 2.1 h (Timmer et al, 2000). The scans were timed to coincide with the time of peak plasma concentration of mirtazapine after an oral dose (emission scan start time 90 mins after ingestion of tablet). All subjects had an indwelling venous cannula inserted into the median cubital vein of the forearm to inject the radioligand and also had an indwelling arterial cannula in the radial artery to withdraw blood for the measurement of the PET input function.

Two out of the five subjects had their blocking scan before the baseline scan, the other three had the tracer alone scan before the scan with mirtazapine pretreatment. The time interval between the two PET scans in individuals ranged between 6 and 83 days. The Research Ethics Committee of the Hammersmith Hospitals Trust and the Administration of Radioactive Substances Advisory Committee (ARSAC) of the United Kingdom approved the study.

Positron Emission Tomography Scans

The radiotracer [¹¹C]MDL 100,907 was synthesized according to the method previously described by Lundkvist et al (1996), and was injected by hand into an antecubital vein as a smooth bolus over 30 secs. The injected radioactivity dose was between 337 and 379 MBq (mean: 361 MBq, SD: 14 MBq). The radiochemical purity of the injected [¹¹C]MDL 100,907 was high and ranged from 97.9% to 100% with a mean of 99.4% and an SD of 0.8%. The injected mass of MDL 100,907 varied between 0.8 and 8.9 μg with a mean value of 3.4 μg and an SD of 2.6 μg. The specific activity was on average 65 GBq μmol⁻¹ with an SD of 48 GBq μmol⁻¹. The dose of MDL 100,907 injected in a subject in one scan ranged from 0.021 to 0.293 nmolkg⁻¹ body weight. In rat brain scans with similar specific activity, Hirani et al (2003) have found no effect of coinjected MDL 100,907 between 0.50 and 0.97 nmol kg⁻¹ on the specific binding of [¹¹C]MDL 100,907.

All PET scans were performed on the high-sensitivity Siemens/CTI scanner ECAT EXACT3D with an axial field of view of 23.4 cm and 95 reconstructed transaxial image planes (Spinks et al, 2000). A 5-mins transmission scan using a ¹³⁷Cs point source was performed before each study for subsequent attenuation correction (Xu et al, 1994) and scatter correction (Watson et al, 1996). The 95-mins three-dimensional (3D) dynamic emission scan was acquired in list mode. In the postacquisition frame rebinning, 28 time frames of increasing length were generated (30 s background frame before the injection, then three 10s-frames, three 20s-frames, three 30s-frames, three 60s-frames, four 120s-frames, six 300s-frames and five 600s-frames). The spatial resolution of the images reconstructed using the reprojection algorithm (Kinahan and Rogers, 1989) with the ramp and Colsher filters set to Nyquist frequency is close to isotropic: 5.1 mm full-width at half-maximum (FWHM) transaxially and 5.9 mm FWHM axially averaged over a radius of 10 cm from the centre of the field of view (Spinks et al, 2000).

Arterial whole-blood activity was monitored continuously for the first 15 mins of the scan with a bismuth germanate coincidence detector (Ranicar et al, 1991). A total of 10 discrete arterial blood samples were taken at 5, 10, 15, 20, 30, 40, 50, 60, 75 and 95 mins into heparinized syringes. The activity concentration of the whole blood and plasma were measured.

Eight plasma samples per scan (at 5, 15, 30, 40, 50, 60, 75 and 95 mins) were analysed for metabolites using a semiautomated system with on-line solid-phase extraction (SPE; small column packed with C-18) followed by reverse-phase chromatography (Luthra et al, 1993). The plasma samples were loaded onto the SPE column and eluted with diammoniumhydrogen phosphate (0.01 mol/L) and further analysed using reverse-phase column (μ–Bondapak C₁₈ column, 30 × 0.78 cm² i.d.) eluted with a mixture of ammonium acetate (10 mmol/L), acetonitrile and trifluoroacetic acid (55:45; 0.1 v/v) at a flowrate of 3 ml/min. In order to identify the peak representing the unmetabolized compound, cold standard MDL 100,907 was added. In one selected case, cold MDL 105,725 was also added such that the position of the peaks of the radiolabelled metabolites relative to MDL 105,725 could be located. The eluate was then monitored for ultraviolet (UV) absorbance at 210 nm, and radioactivity and subsequent data captured on a PC-based integrator. The amount of [¹¹C]MDL 100,907 and its radioactive metabolites at a given time point was calculated from the decay-corrected integrated radiochromatogram and the levels of radioactivity in the SPE eluate and expressed as a percentage of total plasma sample injected.

Magnetic Resonance Scans and Definition of Volumes of Interest

For each subject, brain magnetic resonance images (MRI) were acquired with T₁ weighted RF spoiled gradient echo volume scans on an 1 T Philips Medical Systems HPQ + Scanner. The echo time T_E was 6 ms, the repetition time T_R was 21 ms, the flip angle was 35°, yielding an image resolution of 1.6 mm × 1.6 mm × 1.0 mm (AP, LR, HF, respectively).

A set of volumes of interest (VOI) was defined with a probabilistic brain atlas template (Hammers et al, 2002, 2003). The following 11 VOIs were chosen for the kinetic analysis of [¹¹C]MDL 100,907 (the volume of these VOIs in the defining template is given in parenthesis):

After coregistering the individual MRI to the PET image summed from 7 to 95 mins after tracer injection using an automated multiresolution optimization procedure (Studholme et al, 1997), regional time–activity curves were generated from the dynamic images using the medical imaging software ANALYZE (Robb and Barillot, 1989; Robb et al, 1989).

Data Analysis

For the generation of the plasma input functions, the time course of the plasma-to-blood activity concentration ratio, obtained from the first four discrete arterial samles at 5, 10, 15 and 20 mins scan time, was first fitted to a model function. Then the measurement of the arterial whole-blood activity concentration obtained from the continuous detector system (Ranicar et al, 1991) was multiplied with that model function to obtain a total plasma activity concentration curve for the first 15 mins of the scan. This curve was then combined with the discrete plasma activity concentration measurements at 20, 30, 40, 50, 60, 75 and 95 to obtain an input function describing the total plasma activity concentration for the entire scan. Finally, the input function of the activity concentration due to unmetabolized [¹¹C]MDL 100,907 in plasma was created by multiplying the total plasma activity input function with the function obtained from the fit of the model for the parent fraction in plasma to the eight measurements of the parent compound during the scan. Calculations were performed using MATLAB® (The Math-Works, Inc., Natick, MA, USA) on Sun Ultra™ 10 workstations (Sun Microsystems, Inc., Santa Clara, CA, USA).

Estimates of the compartmental model microparameters were obtained from weighted fits as previously described (Gunn et al, 1998).

In all reversible plasma input function compartmental models, the total volume of distribution VD is given by the integral of the impulse response function (Gunn et al, 2001). With the definition of binding potential BP introduced by Mintun et al (1984), f₁ as the fraction of radioligand not bound in plasma and f₂ as the free fraction of radioligand in tissue, estimates of binding potential in a target region with specific binding can be obtained from the estimated VD in that region and the estimated VD in a reference region without specific binding (Gunn et al, 2001)

f 1 B P = V D target region - V D reference region

f 2 B P = V D target region V D reference region - 1

Watabe et al (2000a) proposed a reference tissue input function model with three compartments for the quantification of dynamic studies with [¹¹C]MDL 100,907. The kinetic behaviour in the reference tissue is described based on two tissue compartments (similar to Figure 6B except that the second tissue compartment is for nonspecific binding) using four rate constants K′₁, K′₂, K′₅ and K′₆. For the target region, one tissue compartment with the rate constants K₁ and k₂ is assumed to characterize the combined kinetics of free, non-specific and specific binding (Figure 6C). Time–activity curves of the target regions were fitted using a cerebellar reference tissue input function and the system impulse response function H_TR(t) for the Watabe reference tissue model as described by Gunn et al (2001):

H T R ( t ) = R I ( δ ( t ) + Φ 1 e - θ 1 t + Φ 2 e - θ 2 t )

with

R I = K 1 K 1 ′ δ ( t )

the symbolic Dirac delta function δ(t)

Φ 1 = k 2 ′ k 5 ′ k 5 ′ + k 6 ′ - k 2 ′ θ 1 = k 5 ′ + k 6 ′

Φ 2 = k 2 2 - k 2 ( k 2 ′ + k 5 ′ + k 6 ′ ) + k 2 ′ k 6 ′ k 5 ′ + k 6 ′ - k 2 θ 2 = k 2

As the integral of the impulse response function of all reference tissue input function models with reversible kinetics in both the target and the reference region is equal to VD^{target region /} VD^{reference region} (Gunn et al, 2001), a binding potential estimate was calculated from:

f 2 B P = R I ( 1 + Φ 1 θ 1 + Φ 2 θ 2 ) - 1

The regional receptor occupancy in a subject was estimated from the ratio of the regional binding potentials in the blocked scan over the regional binding potentials in baseline scan and expressed as a percentage:

O c c u p a n c y = ( 1 - ( f i B P ) blocked scan ( f i B P ) baseline scan ) × 100 % .

Statistical analyses were performed using Statistical Package for Social Sciences SPSS version 11 (SPSS Inc., Chicago, IL, USA). Values are given as mean ± standard deviation (SD) unless specified otherwise. Significant interactions were explored using a two tailed paired samples t-test. All correlations were performed using Pearson's product moment correlation coefficient r.

Input Function

After intravenous injection of the [¹¹C]MDL 100,907 bolus, the radioactivity concentration in whole blood was higher than in plasma throughout the time of the PET scan as reported by Ito et al (1998). During the first phase of the scan (approximately 20 mins), the plasma-to-blood activity ratio decreased further, and after it reached a minimum in the middle of the scan it began to rise (Figure 1). This later rise is indicative of the appearance of radiolabelled polar metabolites, which tend to accumulate in plasma rather than in red cells.

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Figure 1

Ratio of plasma activity concentration over whole-blood activity concentration obtained from the ten PET scans (five subjects, baseline and blocked scans). Depicted are the mean values of the 10 discrete arterial blood samples from each scan connected with the solid line with their standard deviations as bars.

A polynomial was tested as the model function describing the time course of the plasma-over-whole-blood activity concentration ratio (POB ratio) during the first 15 mins of the scan when the continuously operating blood detector system (Ranicar et al, 1991) was used to record the whole-blood activity concentration. Since the decrease of the measured POB ratios over the considered period was small compared with the variability inherent in these POB ratios originating from the division of two radioactivity measurements, a polynomial with order zero was sufficient to describe the data:

POB ( t ) = p 0 f o r t ≤ 15 min

The estimated POB ratios for all five subjects under baseline and blocked condition are given in Table 1. In all subjects a small increase of the parameter p₀ in the blocked scan relative to the baseline scan was observed. This increase was on average only 1.6%, with the biggest increase observed in subject 4 with 2.5%. Ito et al (1998) suggested that [¹¹C]MDL 100,907 may bind to the 5-HT_2A receptors in platelet membranes. The small albeit consistent across subjects increase of the POB ratio observed in this study is in agreement with the possibility that the administration of mirtazapine lead to a blockade of these placelet binding sites, hence reducing the activity concentration in the red cell fraction of the blood sample relative to the activity concentration in the plasma fraction.

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Table 1

Parameter estimates of the model functions for the plasma-over-whole-blood activity concentration ratio (p_o) and for the parent fraction in plasma (q₁, q₂ and q₃). Values are given with their SE on the estimates

Subject	Scan	po (unitless)	q 1 (min −1 )	q 2 (unitless)	q 3 (unitless)
1	Baseline	0.81243 ± 0.00919	0.01603 ± 0.00437	0.51364 ± 0.19343	0.05167 ± 0.01212
	Blocked	0.82423 ± 0.00312	0.03365 ± 0.00956	0.25949 ± 0.06786	0.06579 ± 0.02594
2	Baseline	0.77423 ± 0.01024	0.03733 ± 0.00424	0.16018 ± 0.01614	0.04439 ± 0.00635
	Blocked	0.78991 ± 0.00790	0.02853 ± 0.00848	0.19266 ± 0.05617	0.07369 ± 0.01870
3	Baseline	0.78029 ± 0.00678	0.06956 ± 0.02951	0.14664 ± 0.04003	0.09415 ± 0.03499
	Blocked	0.78687 ± 0.01112	0.03980 ± 0.00958	0.23301 ± 0.05352	0.09196 ± 0.01860
4	Baseline	0.80059 ± 0.00506	0.03803 ± 0.00979	0.39164 ± 0.10715	0.08396 ± 0.02631
	Blocked	0.82125 ± 0.00368	0.08982 ± 0.03421	0.16991 ± 0.03354	0.08628 ± 0.04647
5	Baseline	0.83237 ± 0.00302	0.06419 ± 0.02064	0.26647 ± 0.07172	0.12383 ± 0.04115
	Blocked	0.84370 ± 0.01206	0.12255 ± 0.04283	0.18281 ± 0.02977	0.10285 ± 0.05341
Mean		0.80659	0.05395	0.25165	0.08186
SD		0.02367	0.03260	0.11698	0.02385

The main metabolite of MDL 100,907 at pharmacological doses in rats and dogs has been identified as MDL 105,725 (Scott and Heath, 1998). These authors also reported that MDL 105,725 is able to cross the blood brain barrier (BBB) with a permeability of 25% of the BBB permeability of MDL 100,907. Therefore, the metabolite analysis of a PET scan was done with plasma samples spiked with unlabelled MDL 105,725. As shown in Figure 2, the peak of MDL 105,725 in the UV spectrum does not match with a peak in the radiochromatogram. All radiolabelled metabolites in the radiochromatogram had a shorter retention time on the column than MDL 105,725. They are more polar than MDL 105,725 and much more polar than the parent compound MDL 100,907.

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Figure 2

HPLC analysis of the plasma sample from one subject taken at 15 mins. The solid line depicts the radioactivity measurement, and the dashed line represents the chromatogram from the UV detector. In the UV spectrum, peak 1 corresponds to unlabelled MDL 105,725 and peak 2 comes from the cold standard MDL 100,907. In the radiochromatogram, peak A is some of the unidentified radiolabelled metabolites of [¹¹C]MDL 100,907 and peak B marks the unmetabolized parent compound [¹¹C]MDL 100,907 in plasma.

These findings are in agreement with (Scott and Heath, 1998) who reported that the major phase I metabolite of MDL 100,907 in animals is 3′-O-desmethylated MDL 100,907 (MDL 105,725). Since the [¹¹C]CH3 group in [¹¹C]MDL 100,907 is in that position, the same desmethylation process would result in very polar metabolites labelled with ¹¹C and would leave the MDL 105,725 molecule unlabelled.

The mathematical model for the description of the amount of parent compound in plasma was described by the following equation:

PARENT ( t ) = ( 1 + ( q 1 t ) 2 ) - q 2 - q 3 with q 1 > 0 , q 2 > 0 , q 3 > 0

In Table 1, the estimated parameters of this model function are shown for all five subjects. The original equation using two free parameters q₁ and q₂ was proposed by Watabe et al (2000b). Here, the additional third parameter q₃ reflects the proportion of plasma radioactivity at time zero, which is not due to the parent compound. The estimates of q₃ are based on extrapolated data since no parent fraction measurement could actually be performed before 5 mins scan time. Values of q₃ greater than zero represent co-injected radiochemical impurities or rapidly formed radiolabelled metabolites in the body, for example, created during the first pass of the radioligand from venous blood to arterial blood through the lungs. Since the fraction of radio-chemical impurities measured in the product before injection never exceeded 2.1%, co-injected radiochemical impurities cannot solely explain the estimates of q₃ in Table 1, which range from 0.04439 to 0.12383. The fact that the q₃ estimates of subjects 1, 3, 4 and 5 are well reproduced in the two scans is consistent with the possibility that a substantial proportion of the parameter q₃ accounts for subject-specific rapid initial metabolism of [¹¹C]MDL 100,907 in the body.

The time courses of the plasma-over-blood ratio (Figure 1) as well as of the parent fraction in plasma (Figure 3) exhibit pronounced inter-individual differences as indicated by the standard deviation bars in these figures. Therefore, the use of population based values for the parent fraction and plasma-over-blood ratio instead of individually measured values cannot be recommended. There is an inverse relationship between these two measurements, that is, those subjects with a parent fraction above population average had a plasma-over-blood ratio lower than the population average and vice versa.

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Figure 3

Fraction of unmetabolized parent compound in arterial plasma. The plotted bars are mean values ±SD from all 10 PET scans. The dashed line is the fitted metabolite model to the population mean values, for which the model parameter estimates with standard errors are q₁ = (0.05463 ± 0.00573) min⁻¹, q₂ = 0.19635 ± 0.01490 and q₃ = 0.08991 ± 0.01061. In Table 1, the parameter estimates for all scans are presented.

The difference between the estimated parameters to describe the time course of the parent fraction in plasma for each subject under baseline and blocked condition is smaller than the standard deviation of the population (Table 1). A paired two-tailed t-test of the computed parent fractions at 30 and 60 mins scan time did not reject the null hypothesis that there was no significant difference between the baseline and the blocked scans (P > 0.75). Therefore, no effect of the blocking drug on an alteration of the peripheral metabolism of [¹¹C]MDL 100,907 could be seen. The inter-subject variability (especially of the parameter q₃) is higher than the inter-scan difference for one individual.

Compartmental Analysis

After bolus injection of [¹¹C]MDL 100,907, there was a rapid uptake of radioactivity in brain tissue, as previously described by Lundkvist et al (1996) and Ito et al (1998). There was a clear qualitative difference in the summed PET scans of the baseline and blocked condition for each subject (see Figure 4). In Figure 5A, tissue time–activity curves for a receptor-rich region (occipital lobe), a region with moderate density of 5-HT_2A receptors (thalamus) and the cerebellum are plotted under baseline and blocked condition for subject 1.

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Figure 4

PET images of the mean decay-corrected activity concentration between 7 and 95 mins after blous injection of 370 MBq [¹¹C]MDL 100,907. Sagittal slices of the first subject under baseline condition (top) and in the blocked scan (bottom) are shown. A displacement of the [¹¹C]MDL 100,907 binding in cortical regions is clearly visible whereas no relative change in the cerebellum can be observed.

The regional tissue time–activity curves were initially fitted to the three-tissue compartment, six rate constants model with a free blood volume term using the arterial input function (Figure 6A). The obtained estimates for the rate constants were very sensitive to the choice of the initial parameter values, and the estimated total volumes of distribution had a high coefficient of variation (ratio of the standard error of the parameter estimate over the parameter estimate greater than 30%). Therefore, a two-tissue compartment, four rate constants kinetic model (Figure 6B) was used to obtain estimates of K₁,k₂, k₃, k₄ and fractional blood volume V_b (Kegeles and Mann, 1997). This simplified compartmental structure was introduced by Koeppe et al (1991) for the description of the kinetics of the PET ligand [¹¹C]flumazenil with rapid equilibration between the free and nonspecifically bound tracer pools in brain tissue. Watabe et al (2000b) previously concluded from the analysis of scans in rhesus monkeys that a two-tissue compartment model with four rate constants would be the method of choice among the simplified methods for the analysis of [¹¹C]MDL 100,907 scans. However, it was unclear whether the volume of distribution of the free and nonspecifically bound compartment K₁/k₂ should or should not be fixed to the total volume of distribution obtained in a cerebellar reference region. Since the coefficient of variation of the estimated total volumes of distribution in all brain regions appeared acceptable (< 10%), it was decided to perform the fit of the tissue time–activity curves to the reversible two-tissue compartment model (Figure 6B) with five free parameters (K_1, k₂, k₃, k₄ and V _b ). The K₁/k₂ ratio in the target regions was not constrained to the total volume of distribution of the cerebellum.

In Table 2, a summary of the estimated model parameters of all five baseline scans is presented. The median coefficients of variation for the estimated parameters were 2% for K₁, 21% for k ₂ , 30% for k₃ and 14% for k₄, based on the 11 regional estimates in five subjects. Even in the regions with high binding, that is, the neocortical regions with an average VD greater than 40 (ml plasma) (ml tissue)⁻¹, the coefficients of variation for the k₄ estimates were not significantly higher. The values for the two potential binding potential measures f₁ BP and f₂ BP are given in the last two columns of Table 2. Their between-subject variability is substantially higher than that of VD, which has a mean coefficient of variation of 12%. The mean coefficients of variation derived from the VOIs listed in Table 2 are 30% for f₁ BP and 34% for f₂ BP, respectively. Despite its slightly higher variability, it was decided to choose f₂ BP as the binding potential outcome measure for the calculation of occupancy estimates in the blocked scans, since it was not possible to obtain reliable measurements of the free fraction in plasma f₁ in this study. Therefore, the possibility of changes of f₁ between the baseline and blocked scans could not been eliminated. However, the parameter f₂ is not determined experimentally and the underlying assumption of uniformity of the nondisplaceable compartments can more easily be assessed by a comparison of the reference region VDs. The use of f₂ BP as the preferred outcome parameter is in agreement with previous findings (Laruelle et al, 1994; Carson et al, 1997).

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Table 2

Estimates of the model parameters from the baseline scans (mean values±SD, five normal subjects)

Volume of interest name	K 1 (ml plasma) min −1 (ml tissue) −1	k2 (min−1)	K3 (min−1)	K4 (min−1)	Vb unitless	VD (ml plasma) (ml tissue) −1	f 1 BP (ml plasma) (ml tissue) −1	f 2 BP unitless
Cerebellum	0.379±0.034	0.066±0.014	0.103±0.045	0.045±0.008	0.014 ± 0.009	18.8 ± 1.8
Hippocampus	0.358±0.021	0.043±0.028	0.105±0.091	0.038±0.009	0.033 ± 0.012	32.2±4.4	13.4±5.0	0.73±0.33
Amygdala	0.316±0.035	0.040±0.038	0.108±0.120	0.037±0.009	0.044 ± 0.010	34.4 ± 4.2	15.6±4.3	0.84±0.26
Ant. tem. lobe	0.249 ± 0.023	0.067±0.091	0.208±0.238	0.030±0.014	0.022 ± 0.010	44.3 ± 5.0	25.5±6.1	1.38±0.41
Insula	0.397 ± 0.027	0.044±0.019	0.135±0.082	0.033±0.007	0.032 ± 0.017	46.3 ± 5.8	27.5±5.5	1.47±0.33
Gyrus cing. ant.	0.420 ± 0.058	0.042±0.038	0.155±0.161	0.031±0.006	0.038 ± 0.016	67.4±14.4	48.6±13.9	2.59±0.75
Thalamus	0.490 ± 0.023	0.107±0.116	0.209±0.275	0.051±0.010	0.019 ± 0.020	23.4 ± 1.5	4.6±2.2	0.25±0.14
Occipital lobe	0.448±0.048	0.042±0.015	0.133±0.050	0.032±0.006	0.016 ± 0.014	57.4 ± 6.1	38.6±4.9	2.06±0.23
Putamen	0.493±0.056	0.045±0.010	0.064±0.024	0.048±0.010	0.027 ± 0.015	26.0 ± 2.7	7.2±3.7	0.40±0.23
Frontal lobe	0.393±0.042	0.038±0.008	0.127±0.037	0.034±0.008	0.021±0.014	49.9 ± 5.0	31.1±5.3	1.68±0.38
Parietal lobe	0.390±0.039	0.050±0.028	0.189±0.179	0.035±0.012	0.020 ± 0.014	47.4 ± 7.7	28.6±8.2	1.55±0.54
Mean across VOIs	0.394

A further simplification of the compartmental model to a one-tissue compartment model with two rate constants and fractional blood volume (Figure 6C) significantly decreased the quality of the fit as demonstrated in Figure 5B for three time–activity curves. As previously described (Carson et al, 1998), regions with less specific binding and a smaller VD (e.g. cerebellum, thalamus) appear to require more clearly the two-tissue compartment model than regions with high specific binding (e.g. occipital lobe).

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Figure 5

(A) Three selected regional time–activity curves from the first subject. The symbols on the dashed-dotted lines to the left of the region name refer to the unblocked scan, and the symbols on the dotted lines to the right indicate the blocked scan. (B) Comparison of the fits from the baseline scan of the same subject obtained with the two-tissue compartment model (dashed-dotted lines) and the one-tissue compartment model (dotted line).

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Figure 6

(A) The standard compartmental model used to describe the kinetic behaviour of PET radioligands in brain tissue (Cunningham and Lammertsma 1995) is also referred to as three-tissue compartment, six rate constants model. Two simplifications of this model were introduced by Koeppe et al (1991). (B) The two-tissue compartment, four rate constants model with rapid equilibration between the free and nonspecifically bound compartments. (C) The one-tissue, two rate constants model with rapid equilibration between all tissue compartments with respect to blood–brain barrier transport rates.

Regional Volumes of Distribution, Binding Potentials and Occupancies

Quantification of [¹¹C]MDL 100,907 binding in the brain revealed a wide variation in binding as expected from the preclinical and post-mortem literature. Regional mean VD values of the baseline scans (Table 2) ranged from 18.8 (ml plasma) · (ml tissue)⁻¹ for the cerebellum to 67.4 (ml plasma) · (ml tissue)⁻¹ for the anterior cingulate (anterior cingulated > occipital lobe > frontal lobe > parietal lobe > insula > anterior temporal lobe > amygdala > hippocampus > putamen > thalamus).

There was no significant difference in cerebellar VD between the baseline scan and the mirtazapine scan: VD_baseline= 18.8 ± 1.8 (ml plasma) · (ml tissue)⁻¹, VD_mirtazapine = 17.2 ± 1.6 (ml plasma) · (ml tissue)⁻¹, P> 0.05. However, there was a statistically significant decrease in VD values for all other 10 regions studied (see Figure 7A). A post hoc t-test showed that f₂ BP was significantly reduced across all ten regions after administration of mirtazapine (see Figure 7B). Mean regional VOI occupancy values ranged from 27% in the putamen to 70% in the occipital lobe with the mean occupancy across all VOIs being 60% (see Figure 7C). There appeared to be an age-related decline in binding potential, however, no effect of age on occupancy.

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Figure 7

Comparison of the regional volumes of distribution VD (A) and binding potentials f₂BP (B) between the baseline and the blocked scans. The estimated occupancies (C) are fairly homogeneous across the considered 10 regions of the brain with varying concentrations of 5-HT_2A receptors. The values shown are means with standard deviations obtained from the cohort of five normal subjects, the abbreviations used for the volumes of interest are: cer = cerebellum, hip = hippocampus, amy = amygdala, atl = anterior medial temporal lobe, ins = insula, gca = anterior cingulate gyrus, tha = thalamus, occ = occipital lobe, put = putamen, fro = frontal lobe, par = parietal lobe.

The use of the cerebellar time–activity curve as a reference tissue input function for the reference tissue model with three compartments (Watabe et al, 2000a) did not provide reliable estimates of the five mathematically identifiable microparameters of the model (results not shown). The estimates of the macroparatmeter VD^{target region}/VD^{reference region} of the reversible reference tissue model were generally associated with high standard errors, often exceeding the parameter estimate itself. In consequence, the reference tissue input model analysis did not provide occupancy estimates with a consistency across regions similar to those shown in Figure 7C.