Gating of the neuroendocrine stress responses by stressor salience in early lactating female rats is independent of infralimbic cortex activation and plasticity

<p>In early lactation (EL), stressor salience modulates neuroendocrine stress responses, but it is unclear whether this persists throughout lactation and which neural structures are implicated. We hypothesized that this process is specific to EL and that the infralimbic (IL) medial prefrontal cortex (mPFC) might provide a critical link between assessment of threat and activation of the hypothalamo-pituitary-adrenal (HPA) axis in EL. We measured neuroendocrine responses and neuronal Fos induction to a salient (predator odor) or non-salient (tail pinch) psychogenic stressor in EL and late lactation (LL) females. We found that EL females exhibited a large response to predator stress only in the presence of pups, while responses to tail pinch were reduced independently of pup presence. In LL, HPA axis responses were independent of pup presence for both stressors and only responses to tail pinch were modestly reduced compared to virgins. Intracerebral injection of the local anesthetic bupivacaine (BUP) (0.75%; 0.5 µl/side) in the IL mPFC did not differentially affect neuroendocrine responses to predator odor in virgin and EL females, suggesting that lactation-induced changes in this structure might not regulate stressor salience for the HPA axis. However, the IL mPFC displayed morphological changes in lactation, with significant increases in dendritic spine numbers and density in EL compared to LL and virgin females. EL females also showed improved performance in the attention set-shifting task (AST), which could reflect early plasticity in the IL mPFC at a time when rapid adaptation of the maternal brain is necessary for pup survival.</p

Cell proliferation.

<p>DG proliferation assessed from Ki67-immunostained brain sections from juvenile (P15) and adolescent (P41) male offspring exposed to saline (controls) or nicotine throughout prenatal and postnatal development. Representative micrographs of Ki67-IR cells (arrows) in the subgranular zone or adjacent granule cell layer (gcl) in control P15 (<b>A</b>) and P41 (<b>B</b>) offspring. No significant difference in numbers of Ki67-IR cells was found in the dorsal hippocampus of P15 (<b>C</b>; p = 0.70) or P41 (<b>D</b>; p = 0.17) offspring. Scale bar = 25 µm.</p

Cell survival.

<p>Developmental nicotine exposure did not affect survival of DG cells (p = 0.22). Survival of newborn cells was estimated from BrdU-immunostained brain sections in adolescent (P41) offspring exposed to saline (controls) or nicotine from early embryogenesis until weaning and injected with BrdU at P15.</p

Plasma corticosterone in pups.

<p>Basal plasma corticosterone measured in juvenile pups exposed through breastfeeding to nicotine or saline (controls) from early embryogenesis. Corticosterone levels did not differ between groups (p = 0.67).</p

Neuronal differentiation.

<p>(<b>A</b>) Proportion of BrdU-IR cells differentiating into neurons in adolescent (P41) offspring exposed to saline (controls) or nicotine from early embryogenesis until weaning and injected with BrdU at P15 did not differ between groups (p = 0.48). (<b>B</b>) Orthogonal confocal image of a BrdU/NeuN-labeled DG cell (red: BrdU; green: NeuN). Scale bar = 10 µm.</p

Long-term potentiation.

<p>Long-term potentiation was enhanced by nicotine exposure. Upper panel: Scattered plots of fEPSP against time revealed changes in fEPSP after long-term potentiation (LTP) induction by theta-burst stimulation. Note that LTP triggered in pups exposed to nicotine was stronger than that in control pups. Lower panel: Average traces of fEPSP obtained at time <i>a</i> (before LTP induction) and <i>b</i> (60 min after LTP induction) in representative recordings obtained from saline- and nicotine-exposed pups (% potentiation at 60 minutes after TBS: p = 0.037).</p

Plasma nicotine levels in nicotine-exposed dams and their breastfed pups.

<p>Dam nicotine levels increased non-significantly between pregnancy and post-parturition (p = 0.14). E15: embryonic day 15; P7: postnatal day 7 days post-parturition.</p

Offspring Weight.

<p>Between postnatal day 2 (P2) and weaning (P21), the average weight of male pups exposed to maternal nicotine did not differ significantly from that of saline-exposed controls (ps = 0.16–0.94).</p

Basal synaptic transmission.

<p>Basal excitatory synaptic transmission was not affected by nicotine exposure. Upper panel: averaged traces of evoked field excitatory postsynaptic potential (fEPSP) recorded from the DG of P15 rat pups. The small depolarization ahead of fEPSP is the fiber volley, which represents the amount of activated presynaptic fibers. Basal excitatory synaptic transmission is therefore related to the ratio of fiber volley vs fEPSP at different stimulating currents (see the four averaged traces). We compared this ratio between control and nicotine-treated pups and found no differences between these two groups (lower panel: comparing slope of regression lines fitting fEPSPs and fiber volley amplitudes between two groups, p = 0.21).</p