With up to 6.2% of pregnancies influenced by SSRIs (2
), it is crucial to investigate the consequences of fetal SSRI exposure. To our knowledge, this is the first study that investigated the long term cardiovascular effects from analogous neonatal sertraline exposure. Because the majority of prior animal studies have focused on behavioral aspects, this is a novel area with no preclinical studies for comparison. Likewise, SSRI therapy was only introduced to obstetrical care over the last decade and there are no data on adult cardiovascular phenotypes in exposed humans. The key findings in our model were increased heart rates, increased urinary noradrenaline excretion, and decreased stroke volumes. The cause-effect relationship between the increased heart rates and decreased stroke volumes is not clear, but we hypothesize it may be from increased sympathetic tone, altered vascular capacitance, and/or decreased cardiomyocyte proliferation.
One hypothesis is that the increased heart rates are related to increased central sympathetic outflow. The increased urinary noradrenaline excretion, in combination with increased heart rates and increased activity levels, supports this hypothesis. The increased expression of 5-HTT in the cortex and midbrain demonstrates a rebound increase in the serotonin transporter protein in adults following neonatal 5-HTT inhibition during SSRI exposure. This increase in 5-HTT expression may lead to decreased serotonergic tone and increased sympathetic activation. Heart rate variability (HRV) analysis was developed to assess sympathetic and parasympathetic influences in humans. There is a large body of evidence to suggest that autonomic imbalance, in which the sympathetic system is hyperactive and the parasympathetic system is hypoactive (or increased low frequency: high frequency ratio), is associated with an increased risk of cardiovascular disease (26
). Sertraline exposed mice had increased very low frequency spectral power, but the role of very low frequency power is not entirely understood. In addition, if our findings were entirely related to sympathetic tone then we would have observed significant differences in our heart rate responses following sympathetic blockade. One way to further analyze and support the hypothesis that sertraline exposed mice have increased sympathetic activation would be to measure renal nerve sympathetic activity in these mice.
An alternative hypothesis for increased heart rates and decreased stroke volumes is altered vascular capacitance. Sertraline exposed mice had exaggerated responses to prazosin, an α-antagonist, suggesting altered vascular capacitance. With no other structural differences noted by echocardiograms and no differences noted on hematoxylin-eosin staining, the increased heart rates observed in our mice may be purely related to decreased passive filling. Serotonin’s effects on vascular tone are mediated by multiple receptors which are located on vascular smooth muscle cells. Ungvari et al. demonstrated that fluoxetine elicited substantial dilation of isolated skeletal muscle arterioles and small cerebral arteries of rats (27
) and it is possible early life SSRI exposure permanently altered vascular tone as a result of altered gene expression.
In addition, recent studies have demonstrated a direct effect of serotonin on the developing heart. The 5-HT2B
receptor is expressed in cardiovascular tissues and is the only serotonergic receptor known to be involved in cardiac proliferating cells (28
null mice have heart defects with high mid-gestation lethality (29
). Newborn 5-HT2B
knockout mice develop cardiac dilation with cardiac hypoplasia resulting from impaired proliferation and a decrease in cardiomyocyte size (29
). Although no differences were detected on H&E staining or in overall left ventricular mass, sertraline exposed mice had smaller left ventricular internal diameters in diastole and smaller stroke volumes. Our PCR data demonstrate decreased expression of 5-HT2B
receptors in the sertraline exposed adult mice, suggesting impaired central serotonergic tone. Further studies on isolated neonatal cardiac myocytes are needed to assess impaired growth or proliferation as an etiology of the diminished stroke volumes. Further studies with echocardiograms at earlier time points will also be crucial to determine the onset and trajectory of the diminished stroke volumes. These studies must focus on changes in neonatal mice during a time of rapid cardiomyocyte development that establishes adult myocyte endowment.
The hyperactivity we measured could be secondary to increased sympathetic tone, but is a well described finding in SSRI exposure models (15
). Zeskind et al. also found newborns exposed to intrauterine SSRIs to have higher activity levels (33
). The etiology of this increased locomotor activity is currently an area of active investigation. SSRI exposure may trigger region specific changes in serotonin production or receptor density which could influence physical activity (34
). Importantly, the increased heart rates in sertraline exposed mice were not solely due to activity. When we compared heart rates at specific activity levels, the sertraline exposed mice continued to have higher heart rates than control mice. This hyperdynamic, hyperactive state is consistent with the post-SSRI hypermetabolic state we recently described in identically exposed male and female mice (32
It is important to emphasize why we utilized a neonatal exposure model in mice to mimic late intrauterine SSRI exposure in humans. We were interested in this window since up to 6% of infants are exposed to SSRIs during fetal development (2
). Neonates are rarely prescribed SSRIs and there are typically undetectable SSRI levels in breast milk. In humans, the embryonic stage of cardiac development is complete and the heart has taken its adult shape by the end of the first trimester. The subsequent fetal stage of cardiac development largely establishes the adult complement of myocytes, given that mononucleated cardiomyocytes begin to terminally differentiate into binucleated cells (36
). At the time of delivery, more than 70% of human cardiomyocytes have withdrawn from the cell cycle and the myocyte turnover rate falls to less than 1% per year (37
). Postnatal mice still undergo growth and proliferation of cardiomyocytes during the first 14 days of life (24
). To capture the terminal proliferative phase of cardiomyocyte development without eliciting teratogenic effects from embryonic exposure, we thus utilized a targeted neonatal exposure which captures a window of susceptibility analogous to third trimester in humans. For example, Noorlander et. al demonstrated the teratogenic effects of earlier exposure as mice exposed to fluoxetine during intrauterine phase of cardiac development had significantly decreased survival during the preweaning period (23
). Eighty-one percent of mice treated prenatally with fluoxetine died and were found to have dilated cardiomyopathy (23
), thus eliminating the use of combined intrauterine and neonatal exposure for our experiments.
In our model, we injected mice during the first 14 days and the sertraline dose we used led to levels comparable to typical human levels (38
). Further supporting the biologic relevance of our model, we noted mild neonatal growth restriction, which is consistent with the lower birth weights seen in SSRI exposed infants (4
). Regarding the clinical relevance of the 5 mg/kg dose of sertraline we utilized, we have previously reported plasma levels 2 hours after injection of 71.8 + 1.3 ng/mL and plasma levels 12 hours after injection of 13.1 + 0.6 ng/mL (32
). Based on these levels, there was an estimated peak concentration of 101 ng/mL, and estimated trough concentration of 1.7 ng/mL. Our projected peak concentration approximates that seen in pregnancy, and our projected trough approximates umbilical cord levels (32
). Although our drug levels were comparable to maternal levels during pregnancy, the placenta provides a partial barrier to fetal exposure. It is therefore possible that the lowest dose we utilized exceeds clinical exposure, and the relatively high exposure seen in our model may explain the lack of dose-response.
A major caveat to our study is the lack of clinical correlates. Although multiple studies have been done to determine if SSRIs are cardiac teratogens, no clinical studies have looked at long term cardiovascular effects from neonatal exposure. Our findings of exaggerated hemodynamic responses to sympathetic blockade that suggest increased sympathetic tone could be reinforced by measuring renal nerve sympathetic activity. We only investigated male mice, but previous studies suggest similar phenotypes may be present in female mice.