Interactions between epinephrine, ascending vagal fibers, and central noradrenergic systems in modulating memory for emotionally arousing events

Abstract

It is well-established that exposure to emotionally laden events initiates secretion of the arousal-related hormone epinephrine in the periphery. These neuroendocrine changes and the subsequent increase in peripheral physiological output play an integral role in modulating brain systems involved in memory formation. The impermeability of the blood brain barrier to epinephrine represents an important obstacle in understanding how peripheral hormones initiate neurochemical changes in the brain that lead to effective memory formation. This obstacle necessitated the identity of a putative pathway capable of conveying physiological changes produced by epinephrine to limbic structures that incorporate arousal and affect related information into memory. A major theme of the proposed studies is that ascending fibers of the vagus nerve may represent such a mechanism. This hypothesis was tested by evaluating the contribution of ascending vagal fibers in modulating memory for responses learned under behavioral conditions that produce emotional arousal by manipulating appetitive stimuli. A combination of electrophysiological recording of vagal afferent fibers and in vivo microdialysis was employed in a second study to simultaneously assess how elevations in peripheral levels of epinephrine affect vagal nerve discharge and the subsequent potentiation of norepinephrine release in the basolateral amygdala. The final study used double immunohistochemistry labeling of c-fos and dopamine beta hydroxylase (DBH), the enzyme for norepinephrine synthesis to determine if epinephrine administration alone or stimulation of the vagus nerve at an intensity identical to that which improved memory in Experiment 1 produces similar patterns of neuronal activity in brain areas involved in processing memory for emotional events. Findings emerging from this collection of studies establish the importance of ascending fibers of the vagus nerve as an essential pathway for conveying the peripheral consequences of physiological arousal on brain systems that encode new information into memory storage.

Keywords: amygdala; emotional arousal; epinephrine; learning; memory; vagus nerve.

Figure 1

Time line for the training and testing regimen in the straight alley learning task and dimensions of the training apparatus. Illustration of the electrode implanted along the vagus nerve to produce vagal nerve stimulation.

Figure 2

The runway latencies of each training group during pre-surgical training (A), post-surgical training (B) and during the three days of retention testing (C). There were no significant differences between groups during the training periods. The reduction in reward magnitude from 10 pellets to only 1 pellet on the day of the Shift resulted in significantly longer latencies in the two shifted groups (Sham-Shift and VNS-Shift) relative to the Non-Shifted controls (Sham-NS and VNS-NS). The VNS-Shift group received vagal stimulation after the six training trials with the reduced reward whereas the Sham-Shift group was connected to the stimulator but no current was applied. Retention for the frustrating experience of reward reduction produced by the Shift was assessed one-week later on three daily retention tests. Animals in the vagal stimulation Shift group displayed enhanced retention on the 7 day delayed retention test as evidenced by their continued long latencies to traverse the maze to consume the reduced reward of 1 pellet. Details included in the text.

Figure 3

The effects of systemically administered saline, epinephrine (0.3 mg/kg), sotalol (2.0 or 4.0 mg/kg) or the combination of epinephrine and sotalol on vagal nerve firing in Millivolts. Animals received sotalol 15 min prior to minute (0) and epinephrine at this time point. The first injection at 0 min. Two of the epinephrine injected groups received a second injection at 60 min that consisted of sotalol at 2 mg/kg or 4 mg/kg. One of the saline injected groups received a second injection at 60 min of sotalol at 4 mg/kg. The EPI, EPI + SOT2, and EPI + SOT4 groups exhibited significant increases in vagal firing discharge above baseline for 10–60 min post-epinephrine injection. For the remaining time of recording, vagal activity in the EPI and EPI + SOT2 groups gradually decreased to approximately 20% above basal values. In contrast, vagal activity in the EPI + SOT4 group dropped steeply at 40 min post-sotalol injectionand was reduced by approximately 50% relative to baseline at the end of recording. Neural activity recorded in the form of Millivolts from the vagus nerve in the SAL, SAL + SOT4, SOT2 + EPI, and SOT2 + SAL groups decreased over time with a reduction of approximately 50% relative to baseline at the end.

Figure 4

The effects of systemically administered saline, epinephrine (0.3 mg/kg), sotalol (2.0 or 4.0 mg/kg), or the combination of epinephrine and sotalol on norepinephrine release in the amygdala. Basal levels of norepinephrine were similar across treatment groups during the 60 min drug-free baseline period. Norepinephrine concentrations sampled from the amygdala of the EPI and EPI + SOT2 groups showed consistent and sustained increases throughout the collection period whereas the norepinephrine levels of EPI + SOT4 became comparable to control groups at 180 min post-epinephrine injection. Blocking peripheral beta-adrenergic receptors with sotalol blocked epinephrine induced augmentation of norepinephrine output in the amygdala.

Figure 5

Electrical stimulation of the left vagus nerve significantly increased Fos-like immunoreactivity in each of the examined brain regions bilaterally: NAC, CEA, BLA, HIP, LC, VLM and NTS. In addition, the stimulation also induced an ipsilateral predominance of Fos expression in the LC and NTS. (^*, ^**, and ^*** denotes significant difference between the compared groups with a p-value less than 0.05, 0.01, and 0.001, respectively). Photomicrographs of Fos labeling in the NAC (A), HIP (B), CEA (C), BLA (D), LC (E), VLM (F), and NTS (G) following sham or vagus nerve stimulation. Scale bar = 200 um. (l.v.: lateral ventricle; a.c.: anterior commisure; shell: the shell region of the nucleus accumbens; core: the core region of the nucleus accumbens; 4^th v.: the fourth ventricle; a.p.: area postrema.

Figure 6

Left vagus nerve stimulation activated noradrenergic cells in bilateral LC, bilateral VLM, and ipsilateral NTS. (^*, ^**, and ^*** denotes significant difference between the compared groups with a p-value less than 0.05, 0.01, and 0.001, respectively). Photomicrographs of Fos/DBH labeling in the LC (A), VLM (B), NTS (C,D) following sham or vagus nerve stimulation. Scale bar = 200 um. (4^th v.: the fourth ventricle).

Figure 7

Systemic injection of epinephrine induced significantly more Fos-like immunoreactivity in the NAC, CEA, BLA, HIP, LC, VLM, AP, and NTS. Because injection was given systemically and no hemisphere difference was found, Fos counts from both hemisphere were collapsed. (^*, ^**, and ^*** denotes significant difference between the compared groups with a p-value less than 0.05, 0.01, and 0.001, respectively). Photomicrographs of Fos labeling in the NAC (A), HIP (B), CEA (C), BLA (D), LC (E), VLM (F), and NTS (G) following saline, 0.1 mg/kg or 0.5 mg/kg epinephrine injection. Scale bar = 200 um (a.c.: anterior commisure; shell: the shell region of the nucleus accumbens; core: the core region of the nucleus accumbens; 4^th v.: the fourth ventricle; a.p.: area postrema).

Figure 8

Systemic injection of epinephrine activated noradrenergic cells in the LC, VLM, and NTS. (^*, ^**, and ^*** denotes significant difference between the compared groups with a p-value less than 0.05, 0.01, and 0.001, respectively). Photomicrographs of Fos/DBH labeling in the LC (A), VLM (B), NTS (C,D) following saline or epinephrine injection (0.1 mg/kg). Scale bar = 200 um. (4^th v.: the fourth ventricle).

Figure 9

(A) Percentage of noradrenergic of cell activation. Vagus nerve stimulation induces noradrenergic activation that accounts for 44% of the neuronal activity in the LC, 56% in the VLM, and 8.2% in the NTS. In addition, 0.1 mg/kg epinephrine injection induces noradrenergic activation that accounts for 25% of the neuronal activity in the LC, 39% in the VLM, and 9.6% in the NTS. Moreover, 24% of the Fos activation in the LC, 33% in the VLM, and 7.1% in the NTS are attributed to activated noradrenergic cells induced by 0.5 mg/kg epinephrine. (B) Activated noradrenergic cell distribution. In the vagus nerve stimulation group, out of the total double labeled cells of the three noradrenergic nuclei, 34% was contributed by the LC, 55% by the VLM, and 11% by the NTS. In the epinephrine 0.1 mg/kg group, 20% was contributed by the LC, 47% by the VLM, and 33% by the NTS. In the epinephrine 0.5 mg/kg group, 24% was contributed by the LC, 47% by the VLM, and 29% by the NTS. (C) A numerical summary of the different activation patterns induced by vagus nerve stimulation or systemic epinephrine injection. The findings indicate that heightened levels of plasma epinephrine and increased vagal activity are capable of affecting in a similar fashion, the cellular activity of noradrenergic neurons and its output areas. Epinephrine injection activates high levels of NTS noradrenergic neurons whereas the vagus nerve stimulation recruits more LC noradrenergic activity. As a consequence, epinephrine injected animals exhibited more cell activation in NTS noradrenergic output areas (CEA, NAC) and vagus nerve stimulation animals showed more cell activation in LC noradrenergic output areas (HIP, BLA).