Abstract
Pregnancy is associated with sustained increases in ventilation that support maternal and foetal gas exchange, yet the neural mechanisms linking reproductive hormones to respiratory control remain poorly understood. Serotonergic neurons of the medullary raphe are well-positioned to mediate such effects, as they contribute to central chemosensitivity, modulate respiratory networks, and may be sensitive to endocrine signals. This study investigated whether medullary raphe serotonin (5-hydroxytryptamine; 5-HT) neurons represent a hormonally responsive population capable of influencing respiratory output in female mice.
Immunohistochemical analysis identified prolactin receptor (Prlr) immunoreactivity in a subset of 5-HT neurons within the medullary raphe, whereas colocalisation of oestrogen receptor α (ERα) or progesterone receptor (PR) was not detected under the experimental conditions used. These findings suggest a potential anatomical substrate through which lactogenic hormones could modulate respiratory control circuits. To explore the respiratory contribution of medullary raphe serotonergic neurons, targeted optogenetic and chemogenetic manipulations were combined with chronic intrathoracic pressure (ITP) radiotelemetry in freely-behaving female mice. Because the final optogenetic and chemogenetic datasets comprised two and three animals, respectively, the physiological observations were analysed descriptively. During dioestrus, both animals exhibited lower duty-cycle values during optogenetic photostimulation, but comparable responses were not consistently observed across other respiratory variables or stages of the oestrous cycle. In the chemogenetic experiment, all three animals exhibited increases in inspiratory time, inspiratory pressure and respiratory drive following deschloroclozapine (DCZ) administration. However, the direction and magnitude of these changes relative to the corresponding vehicle responses differed among animals, preventing attribution of the observations specifically to the chemogenetic manipulation.
Subsequent ex vivo electrophysiological recordings demonstrated that photostimulation could evoke action-potential firing in mCherry-positive cells, although firing fidelity was variable. These recordings used a shorter stimulation paradigm than that applied in vivo and therefore, did not establish whether neuronal recruitment could be maintained throughout the five-minute in vivo stimulation period. Histological examination demonstrated reporter expression within the targeted medullary raphe region; however, the neurotransmitter phenotype of hM3Dq-expressing cells and functional neuronal responses to DCZ were not validated. Together, the preliminary physiological observations cannot establish whether optogenetic or chemogenetic manipulation of the targeted medullary raphe neurons alters baseline respiratory output under resting, room-air conditions.
This study establishes a methodological framework combining radiotelemetry with cell-specific neuronal manipulation in freely behaving female mice and provides anatomical evidence that a subset of medullary raphe serotonergic neurons has the potential to respond to lactogenic signalling. However, pregnancy and lactogenic signalling were not manipulated directly; therefore, the findings motivate rather than demonstrate a role for these neurons in pregnancy-associated respiratory adaptation. Future studies should use adequately replicated cohorts and validated manipulation protocols to examine these circuits during pregnancy, following direct manipulation of lactogenic signalling, and under controlled respiratory challenge.