Sharmin Maswood

Activation of serotonin-immunoreactive cells in the dorsal raphe nucleus in rats exposed to an uncontrollable stressor.

The dorsal raphe nucleus (DRN) and its serotonergic terminal regions have been suggested to be part of the neural substrate by which exposure to uncontrollable stressors produces poor escape responding and enhanced conditioned fear expression. Such stressor exposure is thought to selectively activate DRN serotonergic neurons in such a way as to render them transiently sensitized to further input. As a result of this sensitized state, behavioral testing procedures are thought to cause excess serotonergic activity in brain regions that control these behaviors. The present studies were conducted to investigate activity in the DRN following exposure to escapable and yoked, inescapable tailshock. Neural activity was characterized using immunohistochemistry to detect the immediate early gene product Fos in serotonin-immunoreactive cells in the DRN. Inescapable tailshock led to greater serotonergic neural activity than did escapable tailshock, supporting the hypothesis that uncontrollable stressors preferentially activate serotonergic neurons in the DRN.

Exposure to inescapable but not escapable shock increases extracellular levels of 5-HT in the dorsal raphe nucleus of the rat

The effects of escapable and yoked inescapable electric tailshocks on extracellular levels of serotonin (5-HT) in the dorsal raphe nucleus were measured by in vivo microdialysis. In comparison to either control rats or to their own preshock baseline, rats exposed to inescapable shock showed an increase in extracellular 5-HT within 25 min of shock initiation, and 5-HT levels continued to rise during the remainder of the shock session. Rats that were exposed to comparable shock treatment, but that were given the opportunity to escape, did not show an increase in 5-HT. Rats that were restrained but not shocked also did not show an increase in 5-HT. These results add further support to suggestions that serotonergic changes occur in the dorsal raphe nucleus during inescapable shock and that such changes may contribute to the behavioral effects of inescapable shock.

Estrous cycle modulation of extracellular serotonin in mediobasal hypothalamus: role of the serotonin transporter and terminal autoreceptors

In vivo microdialysis was used to examine extracellular serotonin (5-HT) in the mediobasal hypothalamus (MBH) of male and female Fischer (CDF-344) rats. Females from the stages of diestrus, proestrus, and estrus were used. Additionally, ovariectomized rats, primed subcutaneously (s.c.) with estradiol benzoate or estradiol benzoate plus progesterone were examined. Extracellular 5-HT in the MBH varied with stage of the estrous cycle and with the light/dark cycle. Proestrous females had the highest microdialysate concentrations of 5-HT during the light portion of the light/dark cycle and lowest concentrations during the dark portion of the cycle. Diestrous females had the highest levels during the dark portion of the cycle, while males and estrous females showed little change between light and dark portions of the cycle. In ovariectomized rats, there was no effect of 2.5 microg or 25 microg estradiol benzoate (s.c.) on extracellular 5-HT; but the addition of 500 microg progesterone, 48 h after estrogen priming, reduced microdialysate 5-HT near the threshold for detection. In intact females and in males, reverse perfusion with 3 microM fluoxetine, a selective serotonin reuptake inhibitor (SSRI), or 2 microM methiothepin, a 5-HT receptor antagonist, increased microdialysate concentrations of 5-HT. Estrous females and males showed nearly a 4-fold increase in microdialysate 5-HT in response to fluoxetine while smaller responses were seen in diestrous and proestrous rats. In contrast, proestrous rats showed the largest response to methiothepin. Estrous females showed a delayed response to methiothepin, but there was no methiothepin-induced increase in extracellular 5-HT in males. These findings are discussed in reference to the suggestion that extracellular 5-HT in the MBH is regulated in a manner that is gender and estrous cycle dependent. The 5-HT terminal autoreceptor may exert a greater role in proestrous females; the serotonin transporter appears to play a more active role in the regulation of extracellular 5-HT in estrous females and in males.

The dorsal raphe nucleus is a site of action mediating the behavioral effects of the benzodiazepine receptor inverse agonist DMCM.

Systemic administration of benzodiazepine receptor inverse agonists leads to behavioral changes similar to those produced by inescapable shock (IS). The dorsal raphe nucleus (DRN) is a critical structure mediating IS effects. The present experiments determined whether the DRN is a site mediating the behavioral changes produced by benzodiazepine receptor inverse agonists. Microinjection of the inverse agonist Methyl 6,7-Dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM) in the region of the DRN produced enhancement of fear conditioning as assessed by the amount of freezing in the presence of shock cues as well as interference with shuttlebox escape learning assessed 24 hr later. Furthermore, lesion of the DRN blocked the effects of systemic DMCM on fear conditioning and escape learning. These data suggest that the DRN is indeed critical in mediating these behavioral consequences of DMCM and further support a role for the DRN in producing the behavioral changes induced by IS.

Gender and estrous cycle effects of the 5-HT1A agonist, 8-OH-DPAT, on hypothalamic serotonin

Effects of the 5-HT1A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT; 0.04, 0.25, or 1.0 mg/kg), on hypothalamic serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), and their ratio were determined in adult male rats and in diestrous, proestrous, and estrous female rats. Consistent with its action at the somatodendritic 5-HT1A autoreceptor, 8-OH-DPAT decreased the 5-HIAA/5-HT ratio, but the decrease was least evident in proestrous females and in males. Similar to hypothalamic tissue, there was also a decline in the 5-HIAA/5-HT ratio in the hippocampus after treatment with 0.25 mg/kg 8-OH-DPAT. When ovariectomized rats were treated with oil or estradiol benzoate followed 48 h later by oil or progesterone, 0.25 mg/kg 8-OH-DPAT produced a decrease in the hypothalamic 5-HIAA/5-HT ratio in every group except those rats treated with progesterone without estrogen priming. Treatment with estradiol benzoate increased hypothalamic 5-HIAA, and both progesterone and 8-OH-DPAT reduced the metabolite to the level of the ovariectomized control. These results suggest that both estrogen and progesterone contribute to an estrous cycle modulation of the 5-HT1A somatodendritic autoreceptor.

Protective actions of the 5-HT2A2C receptor agonist, DOI, on 5-HT1A receptor-mediated inhibition of lordosis behavior

Sexually receptive proestrous rats were infused bilaterally into the ventromedial nucleus of the hypothalamus (VMN) with 200 ng of 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT), 1000 or 2000 ng of 5-methoxy-3-(di-n-propylamino) chroman (5-MEO-DPAC), or 1000 or 2000 ng of serotonin (5-HT) plus or minus the 5-HT2A/2C agonist, (2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCl (DOI). DOI protected against the lordosis-inhibiting effects of 5-HT and each of the 5-HT1A agonists. These results substantiate a prior report that 2000 ng DOI could protect against the lordosis-inhibiting action of 200 ng 8-OH-DPAT and demonstrate that such protection is not unique to a single 5-HT1A agonist. Consequently, these results strengthen evidence that functionally significant interactions occur among 5-HT receptors within the VMN.

Opioid-dependent effects of inescapable shock on escape behavior and conditioned fear responding are mediated by the dorsal raphe nucleus.

Manipulations of the dorsal raphe nucleus (DRN) modulate the behavioral effects of exposure to inescapable shock (IS). Opiate agonists and antagonists also influence the impact of IS, but the role of the DRN in mediating these effects is unknown. The opiate antagonist naltrexone micro-injected into the region of the DRN immediately prior to IS prevented both the escape deficit and the enhancement of fear conditioning that occur 24 h later. Intra-DRN naltrexone administered at the time of later behavioral testing reduced, but did not eliminate, these effects of prior IS. Conversely, the opiate agonist morphine, in combination with a subthreshold number of 20 IS trials, induced an escape deficit and enhanced conditioned fear 24 h later. Microinjections of naltrexone into the dorsolateral periaqueductal gray area did not alter the effects of IS and electrolytic lesions of the DRN prevented the effect of the morphine-20 IS trial combination. The role of opioids in mediating the behavioral effects of IS is discussed.

Hypothalamic infusion of the 5-HT21C agonist, DOI, prevents the inhibitory actions of the 5-HT1A agonist, 8-OH-DPAT, on lordosis behavior

Sexually receptive, intact, proestrous rats were infused bilaterally into the ventromedial nucleus of the hypothalamus (VMN) with 200 ng of the 5-HT1A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), with 2000 ng of the 5-HT2/1C agonist, (+/-)-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCl (DOI), or with both 8-OH-DPAT and DOI. Alone, VMN infusions of 8-OH-DPAT, but not DOI, inhibited lordosis behavior. When 2000 ng DOI was infused simultaneously with 8-OH-DPAT, the inhibitory effects of 8-OH-DPAT were completely abolished. These results suggest that neural sites responsible for the reported facilitatory effects of 5-HT2/1C agonists on lordosis behavior coexist in the VMN with those sites in which 5-HT1A agonists are effective in reducing lordosis behavior. In contrast to the protective action of the 5-HT2/1C receptor agonist following VMN infusion, no protection was seen when both DOI and 8-OH-DPAT were administered intraperitoneally. Thus, the interaction of these two receptor subtypes in the control of lordosis behavior may be different in regions outside the VMN.

The benzodiazepine receptor antagonists flumazenil and CGS8216 block the enhancement of fear conditioning and interference with escape behavior produced by inescapable shock

Prior work has suggested that the mediation of the behavioral effects of inescapable shock (IS) might involve release of an endogenous β-carboline-like ligand at the dorsal raphe nucleus (DRN) that binds to the benzodiazepine (BZ) recognition site on the GABAA complex, thereby disinhibiting the DRN. This was tested by microinjection of the BZ receptor antagonists flumazenil and CGS8216 in the region of the DRN, either before IS or before later behavioral testing. Both compounds blocked subsequent enhancement of fear conditioning and interference with shuttlebox escape when administered before IS, but had no effect when given before testing. In addition, flumazenil did not alter the behavior of escapably shocked subjects.

Estrogen-progesterone and 8-OH-DPAT attenuate the lordosis-inhibiting effects of the 5-HT1A agonist in the VMN

Ovariectomized rats were treated for 2 consecutive weeks with 25 micrograms estradiol benzoate followed 48 h later with 500 micrograms progesterone. Bilateral infusions of 200 ng 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) into the ventromedial nucleus of the hypothalamus (VMN) inhibited female sexual behavior on the first but not the second week of hormone priming. Such attenuation on the second week of priming did not appear to result from an enhanced receptivity of the female rats since there were no differences in the L/M ratios prior to drug infusion; nor was the attenuation a consequence of infusion-induced VMN damage since neither saline nor 8-OH-DPAT preinfusions prevented the later inhibitory effects of 8-OH-DPAT on lordosis behavior. However, preinfusion with 8-OH-DPAT may have reduced the duration of the inhibition. Hormone priming (without any VMN infusion) also partially attenuated the effect of 8-OH-DPAT. Both hormone priming and treatment with 8-OH-DPAT were required to eliminate the effects of the second 8-OH-DPAT treatment. Thus, the present results suggest that gonadal hormones, alone, slightly attenuate the effects of agonist activation of 5-HT1A receptors involved in the inhibition of lordosis behavior; that agonist activation of 5-HT1A receptors also produces a slight attenuation; but that both treatments together have a robust protective action against the inhibitory effect of a 5-HT1A agonist on female lordosis behavior.