PET/fMRI reveals no functional response to 5-HT₄ drugs even at high receptor occupancy

Objectives: The use of simultaneous PET/fMRI allows concurrent measurement of receptor occupancy using PET and neurovascular responses using fMRI in a single scan session. This synergistically combines the molecular specificity of PET and the spatiotemporal resolution of fMRI applicable to a unified PET/fMRI model for estimation of neurotransmitter release profiles. In our previous work we demonstrated the utility of simultaneous PET/fMRI to measure occupancy and neurovascular responses (or changes in cerebral blood volume [CBV]) in the dopaminergic system using D₁- and D₂-receptor drugs, and in the serotonergic system we recently reported initial results from 5-HT_1A and 5-HT_2A receptor specific targeted agonists and antagonists. 5-HT_2A agonists increased CBV whereas 5-HT_1A agonist decreased CBV corresponding to these receptors inherent excitatory and inhibitory properties, respectively. In contrast to agonists which directly stimulate the receptor, antagonist-elicited changes in CBV are indirect and result from blocking the effects of endogenous neurotransmitter. This was observed in the 5-HT_2A and 5-HT_1A receptor antagonist studies where receptor 5-HT_2A and 5-HT_1A subtype specific antagonists caused the opposite responses as their agonist counterparts; the magnitude of antagonist responses was enhanced when endogenous levels of 5-HT were elevated due to the greater endogenous signal available to attenuate. In this work, we extend upon our prior analysis of the 5-HT system to measure the receptor occupancy and functional responses of agonist and antagonist drugs targeting the excitatory 5-HT₄ receptor.

Methods: Two rhesus macaques were scanned a total of 9 times using a simultaneous PET/MR (Siemens Trio with BrainPET insert). Dynamic PET imaging was performed following the bolus injection of the 5-HT₄ radiotracer ¹¹C-SB207145. Baseline scans were acquired using radiotracer only, and competition scans were acquired using a coinjection of ¹¹C-SB207145 plus a competitor drug (either antagonist SB207145 or agonist RS67333). Changes in CBV elicited from 5-HT₄ agonist and antagonist drugs were measured using fMRI with iron-oxide enhanced echo planar imaging and analysis was focused in the striatum. Responses from the 5-HT₄ antagonist drug was also measured under ‘elevated 5-HT’ conditions using a pretreatment of fenfluramine (5mg/kg i.v. infused over 5 minutes 30 minutes prior to antagonist). Elevating 5-HT increased the amount of endogenous 5-HT₄ signaling that could be silenced by the exogenous 5-HT₄ antagonists and thus enhances the drug’s indirect effects. Fenfluramine only scans were also acquired for comparison with the elevated 5-HT scan with antagonist competitor.

Results: 5-HT₄ agonist RS67333 did not elicit changes in CBV even at the highest dose (0.2 mg/kg) that resulted in a 90% 5-HT₄ striatal receptor occupancy. Similarly, the antagonist SB207145 did not elicit a change in CBV even at a dose of 0.25 mg/kg which caused >90% 5-HT₄ receptor occupancy. Moreover, SB207145 did not elicit a change in CBV at 0.25 mg/kg even when endogenous levels of 5-HT were increased using a fenfluramine pretreatment.

Conclusion: Direct and indirect receptor mediated signaling of the 5-HT₄ system by targeted agonist and antagonist drugs resulted in no measureable fMRI changes in CBV. In contrast to the 5-HT_1A and 5-HT_2A receptor subtypes, drugs targeting the 5-HT₄ receptor do not elicit an observable fMRI response. Hence, the 5-HT₄ receptor can potentially be omitted from consideration in the unified PET/fMRI model of pharmacologic action of the serotonergic system. Support: R01MH100350