Research reportExposure to inescapable but not escapable shock increases extracellular levels of 5-HT in the dorsal raphe nucleus of the rat
Introduction
There has been considerable recent interest in the involvement of serotonergic systems in mediating behavioral and endocrine sequelae of exposure to stressors 14, 15, as well as the role of serotonin (5-HT) in stress-related psychological conditions such as anxiety 16, 17, 25and depression 35, 44. Much of this work has focused on ascending 5-HT projections, particularly those from the dorsal raphe nucleus (DRN). Exposure to diverse stressors including immobilization [34], footshock [10]and aggressive encounters [24]have been reported to alter 5-HT metabolism. In addition, forced swimming [23], exposure to a stimulus previously paired with footshock [49], exposure to another rat receiving footshock [22], placement on an elevated plus maze [43], and a saline injection [2]each increase 5-HT efflux in projection regions of the DRN as measured by in vivo microdialysis.
Although a variety of stressors appear to produce 5-HT release in projection regions of the DRN neurons, different stressors produce quite different patterns 2, 24, 42. Furthermore, the same stressor can simultaneously lead to increases in 5-HT efflux, no change, and even decreases in different projection regions of the DRN [23], and some stressors produce little or no change in DRN neuronal activity [48]. It is not known which aspects of a stressor (severity, propensity to produce motor activation, consequent arousal, etc.) might modulate stressor-induced alterations in DRN 5-HT function [42]. Pharmacological and lesion data suggest that the degree to which the organism can exert behavioral control over the stressor might be of importance. Exposure to stressors which are inescapable and unavoidable (uncontrollable) produces a constellation of behavioral changes that are not produced by escapable (controllable) stressors that are exactly equal in intensity, duration, and temporal distribution 3, 26, 47. Inescapable shock (IS) has been the most frequently used uncontrollable stressor, and interference with subsequent escape learning has been the most widely explored behavioral consequence. It is of note here that the interference with escape learning normally produced by IS can be blocked by lesion of the DRN [28]and by microinjection before IS of pharmacological agents into the DRN that inhibit the activity of 5-HT neurons 29, 30. In addition, lesion of the DRN blocks the interference with escape produced by systemic drugs which mimic IS action [27]. Furthermore, intra-DRN microinjection of agents that increase 5-HT activity can produce some of the same effects on later behavior as does IS [27]. These data indicate that activation of 5-HT neurons within the DRN is important in mediating behavioral consequences of IS, consequences which do not follow escapable or controllable shock. This suggests that stressor controllability might be a factor that modulates the impact of stressors on 5-HT activity within the DRN. Consistent with this possibility, Petty et al. [40]reported differences in 5-HT efflux following behavioral escape testing 24 h after IS between rats that did, and did not, demonstrate failure to learn to escape.
DRN 5-HT function has not been investigated during exposure to IS, nor has DRN 5-HT function been compared during equivalent uncontrollable and controllable stressors. The purpose of the present experiment was therefore to begin to assess DRN 5-HT function during IS and during matched escapable shock (ES). An examination of extracellular 5-HT within the DRN, rather than in projection regions of the DRN, was chosen for initial study. High levels of 5-HT can be measured within the DRN. Indeed, basal levels are higher than in projection regions such as the ventral hippocampus [33]. The DRN is extremely rich in 5-HT cell bodies and dendrites, but contains few 5-HT terminals 5, 8, 38. The extracellular 5-HT measured within the DRN is therefore likely to be somatodendritic in origin and may reflect impulse activity in DRN 5-HT neurons [33]. These facts, together with the finding that different projection regions of the DRN can yield completely different patterns of 5-HT release in response to the same stressor [23], suggested the DRN itself as a reasonable starting point.
Section snippets
Materials and methods
Male Sprague–Dawley rats (Harlan labs), 90–100 days old, were maintained on a 12:12 h light–dark cycle with all experimentation occurring during the light portion of the cycle. Experimental procedures were in accordance with protocols approved by the University of Colorado Institutional Animal Care and Use Committee.
Rats were anesthetized with a combination of Ketamine (60 mg/kg) and Xylene (13 mg/kg) and implanted with a single CMA 12 microdialysis probe guide (CMA/Microdialysis, Acton, MA)
Results
One animal from the ES group was found to have a lesion near the site of the probe and was eliminated from the data analysis. For the remaining animals baseline values of 5-HT did not differ across treatment conditions (p>0.05). The mean±S.E. (pg 5-HT/5 μl) for baseline samples was 0.75±0.18, 0.60±0.22, 0.58±0.22 and 0.81±0.14 for C, R, IS and ES groups, respectively. Fig. 1 shows 5-HT levels for each of the groups at the various stages of the experiment. IS produced a clear increase in
Discussion
Previous experiments have measured 5-HT in projection regions of the DRN during and following exposure to a stressor, but have not assessed extracellular 5-HT within the DRN. Indeed, there has been some question whether extracellular 5-HT within the DRN is dependent on nerve impulses of DRN 5-HT neurons, involves vesicular release, or is even under local receptor regulation [1]. However, vesicles are present in DRN 5-HT dendrites [7]and more recent work indicates that extracellular 5-HT within
Acknowledgements
We would like to thank Dr. Jose Amat for developing many of the procedures that allow sampling during exposure to the stressor. This research was supported by MH50479 and MH00314.
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