Role of serotonin in the lack of sensitization caused by prolonged food deprivation in Aplysia

Abstract

Food deprivation may cause neurological dysfunctions including memory impairment. The mollusk Aplysia is a suitable animal model to study prolonged food deprivation-induced memory deficits because it can sustain up to 14 days of food deprivation (14DFD). Sensitization of defensive withdrawal reflexes has been used to illustrate the detrimental effects of 14DFD on memory formation. Under normal feeding conditions (i.e., two days food deprivation, 2DFD), aversive stimuli lead to serotonin (5-HT) release into the hemolymph and neuropil, which mediates sensitization and its cellular correlates including increased excitability of tail sensory neurons (TSNs). Recent studies found that 14DFD prevents both short-term and long-term sensitization, as well as short-term increased excitability of TSNs induced by in vitro aversive training. This study investigated the role of 5-HT in the absence of sensitization and TSN increased excitability under 14DFD. Because 5-HT is synthesized from tryptophan obtained through diet, and its exogeneous application alone induces sensitization and increases TSN excitability, we hypothesized that 1) 5-HT level may be reduced by 14DFD and 2) 5-HT may still induce sensitization and TSN increased excitability in 14DFD animals. Results revealed that 14DFD significantly decreased hemolymph 5-HT level, which may contribute to the lack of sensitization and its cellular correlates, while ganglia 5-HT level was not changed. 5-HT exogenous application induced sensitization in 14DFD Aplysia, albeit smaller than that in 2DFD animals, suggesting that this treatment can only induce partial sensitization in food deprived animals. Under 14DFD, 5-HT increased TSN excitability indistinguishable from that observed under 2DFD. Taken together, these findings characterize 5-HT metabolic deficiency under 14DFD, which may be compensated, at least in part, by 5-HT exogenous application.

Keywords: Excitability; Food deprivation; Sensitization; Serotonin; Tail sensory neuron.

Fig 1.

5-HT concentrations measured from hemolymph samples of 2DFD and 14DFD Aplysia. Representative total ion current (TIC) chromatograms show hemolymph 5-HT in 2DFD (A1) and 14DFD (A2) animals. Black dash line brackets highlight the area where the peak of 5-HT is supposed to occur based on 5-HT standards with known concentrations. (B) Summary data revealed that 5-HT concentration in the hemolymph was significantly reduced in 14DFD animals compared to 2DFD animals.

Fig 2.

5-HT concentrations measured from the ganglia samples of the 2DFD and 14DFD Aplysia. Representative TIC chromatograms show ganglia 5-HT in 2DFD (A1) and 14DFD (A2) animals. Black dash line brackets highlight the area where the peak of 5-HT is supposed to occur based on 5-HT standards with known concentrations. (B). Summary data revealed that in the ganglia there was no significant difference in 5-HT concentrations between 2DFD and 14DFD animals.

Fig. 3.

In vivo exogenous application of 5-HT induced LTS under both 2DFD and 14DFD 24 h after the treatment. (A) TSWR duration was tested 30 min before, and 24 h after 5-HT/ASW treatment, to evaluate the ability of 5-HT to induce LTS. (B) LTS was observed in both 2D-5-HT and 14D-5-HT Aplysia compared to corresponding ASW groups. However, the amount of LTS induced by 5-HT was significantly smaller under 14DFD compared to 2DFD. Shared capitalized letters denote statistical similarity among groups.

Fig. 4.

In vitro bath application of 5-HT induced increased excitability of TSNs in 2DFD and 14DFD preparations both 5 min and 15 min after treatment. (A) TSN properties were measured 10 min prior to, 5-min after and 15-min after treatment with 5-HT/ASW, to analyze change of TSN excitability by 5-HT. (B1-B4) Sample traces of TSN spikes in pre-tests, 5-min and 15-min post-tests in each of the four groups. (C1) Summary data illustrate that increased excitability of TSNs was present both in 14D-5-HT preparations and 2D-5-HT preparations 5 min after in vitro application of 5-HT. (C2) Summary data illustrate that increased excitability of TSNs was observed both in 14D-5-HT preparations and 2D-5-HT preparations 15 min after in vitro application of 5-HT. Shared capitalized letters denote statistical similarity among groups.