Annabell C. Segarra

The neural consequences of repeated cocaine exposure revealed by functional MRI in awake rats

The use of functional magnetic resonance imaging (fMRI) in animal models of cocaine addiction is an invaluable tool for investigating the neuroadaptations that lead to this psychiatric disorder. We used blood-oxygen-level-dependent (BOLD) MRI in awake rats to identify the neuronal circuits affected by repeated cocaine administration. Rats were given an injection of cocaine (15 mg/kg, i.p.) or its vehicle for 7 days, abstained from injections for 1 week, and challenged with an intracerebroventricular cocaine injection during functional imaging. Acute cocaine produced robust positive BOLD responses across well-known monoamine-enriched brain regions, such as the prefrontal cortex, nucleus accumbens, dorsal striatum, sensory cortex, hippocampus, thalamus, and midbrain areas. However, repeated cocaine administration resulted in lower BOLD responses in the prefrontal cortex, agranular insular cortex, nucleus accumbens, ventral pallidum, and dorsomedial thalamus, among other brain regions. Reductions in BOLD intensity were not associated with variations in cerebrovascular reactivity between drug naïve rats and those repeatedly exposed to cocaine. Therefore, the lower metabolic activation in response to cocaine could reflect a reduced neuronal and/or synaptic activity upon repeated administration.

ACTH modulation of nerve development and regeneration

(1) The availability of short amino acid sequences of the naturally occurring ACTH 1-39 molecule has made it possible to separate the corticotropic characteristics of the parent molecule from its neurotrophic effects. Potent neurotrophic fragments are ACTH 4-10, an analog of ACTH 4-9 (Org 2766), and alpha-MSH (ACTH 1-13), peptide fragments that do not evoke corticosteroid secretion, yet clearly affect both the development and regeneration of peripheral nerve. (2) Early postnatal administration of either ACTH 4-10 or Org 2766 accelerates the neuromuscular development of the immature rat, increasing the contractile strength of the EDL muscle and inducing more rapid muscle contractions. Grasping strength and motor activity are increased; these are all changes indicative of more rapid neuromuscular maturation. Prenatal peptide treatment elicits a more complex pattern of response since administration early in gestation (GD 3-12) accelerates neuromuscular development whereas later administration (GD 13-21) decelerates maturation. (3) ACTH peptides have a similar accelerating effect on the morphology of the developing neuromuscular junction. At two weeks of age, nerve arborization is conspicuously increased by postnatal administration of either ACTH 4-10 or Org 2766, as is nerve terminal branching within the endplate itself. However, this is preceded by an initial depression of nerve branching in the 7-day-old rat pup. We conclude that while the developing neuromuscular system is sensitive to ACTH peptides, this susceptibility is age-related. The crucial role of these peptides may be limited to very brief, defined periods during which the peptides may interact with trophic or growth-associated substances, each of which may have its own decisive, circumscribed time frame of influence. (4) Perinatal administration of ACTH peptides affects CNS development. One measurable indication of this is an acceleration of eye opening. Early exposure to ACTH peptides has long-lasting effects on behavior, apparent when these animals are tested as adults. Increased spontaneous motor activity, heightened states of arousal and agitation, and changes in social behavior have been reported. Certain avoidance responses and tests of visual discrimination in male rats are improved by neonatal treatment with alpha-MSH. Overall motor activity is increased and the normal period of hyperactivity is initiated earlier. Male sexual behavior is decreased and sexually dimorphic behaviors in males are eliminated. alpha-MSH may alter the development of its own dopaminergic feedback circuitry while ACTH affects serotonin levels in the preoptic nucleus.(ABSTRACT TRUNCATED AT 400 WORDS)

Sex differences in models of temporal lobe epilepsy: role of testosterone.

Kainic acid and pilocarpine were used to assess sex differences in temporal lobe seizures. Adult Sprague-Dawley rats were injected with kainic acid (10-12 mg/kg) or with pilocarpine (380 mg/kg) and behavior was recorded for the next 3 h. Trunk blood was collected for hormonal measurements. Our data indicate that the male is more susceptible to the convulsant effects of agents that produce temporal lobe-like seizures. Males presented a higher amount of full limbic convulsions than females. To assess the role of plasma testosterone levels in kainate-induced seizures, a group of males was gonadectomized and half received testosterone replacement. The presence of testosterone, in intact and in gonadectomized males with testosterone replacement, increased the susceptibility to seizure. Seizures were either stronger (full limbic) or more frequent in animals with testosterone compared to animals devoid of testosterone. These results suggest that differences in plasma levels of testosterone may be partially responsible for the observed gender differences in seizure susceptibility. Our data reveal a reciprocal relationship between kainic acid-induced temporal lobe seizures and plasma testosterone. Testosterone enhances the occurrence and the severity of seizures. Conversely, kainic-acid-induced seizures decrease plasma testosterone. The higher plasma corticosterone levels found in these males suggest that kainic acid-induced seizures activate the hypothalamic-pituitary-adrenal axis which may induce alterations in plasma levels of male reproductive hormones.

Imaging cocaine-induced changes in the mesocorticolimbic dopaminergic system of conscious rats

Functional magnetic resonance imaging (fMRI) was used to assess the effects of cocaine on brain activation in fully conscious rats. Methods were developed to image cocaine-induced changes in blood-oxygen-level-dependent (BOLD) signal without the peripheral cardiac and respiratory complications associated with psychostimulant administration. Using spin echo planar imaging (EPI), conscious rats were imaged in a 4.7 T spectrometer prior to and following the intracerebroventricular injection of cocaine (20 microg) in artificial cerebrospinal fluid (10 uL). Within 5 min of injection, there was a significant increase in BOLD signal intensity in the substantia nigra, ventral tegmental area, nucleus accumbens, dorsal striatum and prefrontal cortex, as compared to vehicle controls. Minimal negative BOLD signal changes were observed in response to cocaine and no significant perturbations in normal cardiovascular and respiratory function. These findings demonstrate the technical feasibility of studying psychostimulant-induced brain activity using functional MRI in conscious rats.

Estrogen Increases Spine Density in Ventromedial Hypothalamic Neurons of Peripubertal Rats

Using the single-section Golgi impregnation technique, sex differences in hypothalamic ventromedial (VMN) neurons of gonadectomized juvenile and peripubertal rats were assessed. The effect of estrogen treatment on VMN neurons was also investigated. Juvenile rats were gonadectomized at 16 days of age and peripubertal rats at 36 days. At 5 days after surgery, the rats were injected with estradiol benzoate (20 micrograms/kg) or sesame oil for 2 days after which they were perfused and the brains processed for Golgi impregnation. Estradiol benzoate treatment significantly increased dendritic and soma spine density in juvenile and peripubertal male and female rats. A sex difference was observed in oil- and in estradiol-treated rats, with females exhibiting higher dendritic spine density and a greater response to estrogen priming. Moreover, dendritic and soma spine density was significantly higher in juvenile versus peripubertal rats. It is possible that the sex difference observed in dendritic and soma spine density and in the response to estradiol benzoate treatment is due to an organizational effect of sex steroids.

Estradiol: A key biological substrate mediating the response to cocaine in female rats

A consistent finding in drug abuse research is that males and females show differences in their response to drugs of abuse. In women, increased plasma estradiol is associated with increased vulnerability to the psychostimulant and reinforcing effects of drugs of abuse. Our laboratory has focused on the role of estradiol in modulating the response to cocaine. We have seen that ovariectomy increases the locomotor response to a single cocaine injection, whereas estradiol exacerbates the locomotor response to repeated cocaine administration. Cocaine-induced sensitization of brain activity, as measured by fMRI, is also dependent on plasma estradiol. Moreover, we observed that although all ovariectomized rats show conditioned place preference to cocaine, it is more robust in ovariectomized rats with estradiol. Opioid receptors are enriched in brain regions associated with pleasure and reward. We find that in females, the effectiveness of kappa opioid agonists in decreasing the locomotor response to repeated cocaine varies with plasma estradiol. We also find that estradiol regulates the density of mu opioid receptors in brains areas associated with reward. These data hint that in females, estradiol modulates the behavioral effects of cocaine by regulating mu and kappa opioid signaling in mesocorticolimbic brain structures. Identifying the mechanisms that mediate differences in vulnerability to drugs of abuse may lead to effective therapeutic strategies for the treatment and prevention of addiction and relapse. We encourage health practitioners treating persons addicted to drugs to consider gender differences in response to particular pharmacotherapies, as well the sex steroid milieu of the patient.

Blocking EphA4 upregulation after spinal cord injury results in enhanced chronic pain

Spinal cord injury (SCI) is characterized by a total or partial loss of motor and sensory functions due to the inability of neurons to regenerate. This lack of axonal regenerative response has been associated with the induction of inhibitory proteins for regeneration, such as the Eph receptor tyrosine kinases. One member of this family, the EphA4 receptor, coordinates appropriate corticospinal fibers projections during early development and is expressed in spinal commissural interneurons. Its mechanism of action is mediated by repulsive activity after ligand binding, but its role after trauma is unknown. We examined the temporal expression profile of this receptor after spinal cord contusion in adult rats by RT-PCR and immunohistochemistry. SCI induced a biphasic gene expression profile with an initial downregulation at 2 and 4 days post-injury (DPI) followed by a subsequent upregulation. Double labeling studies localized EphA4 immunoreactivity in neurons from the gray matter and astrocytes of the white matter. To test the role of this receptor, we reduced gene upregulation by intrathecal/subdural infusion of EphA4-antisense oligodeoxynucleotide (ODN) and subsequently assessed behavioral outcomes. No locomotor recovery was observed in the rats treated with the EphA4-antisense ODN. Interestingly, reducing EphA4 expression increased mechanical allodynia, as observed by the Von Frey test and decreased exploratory locomotor activity. These results indicate that upregulation of EphA4 receptor after trauma may prevent the development of abnormal pain syndromes and could potentially be exploited as a preventive analgesic mediator to chronic neuropathic pain.

Perinatal administration of nicotine alters subsequent sexual behavior and testosterone levels of male rats

The effect of pre- and/or postnatal administration of nicotine (0.25 mg/kg) on sexual behavior and testosterone levels in adult male rats was examined. Prenatal nicotine decreases male sexual behavior as measured by the number of males that mounted, intromitted or ejaculated. The males that did mount and/or ejaculate exhibited an increase in mount latency and number of mounts and a decrease in efficiency. This decrease was correlated with a decrease in plasma testosterone levels. Postnatal nicotine treatment improved the sexual performance of sexually naive males as measured by an increase in the number of males that completed 2 ejaculatory series and by a decrease in mount latency. This effect is transient, subsequent testing of the nicotine males eliminated this difference in sexual performance. Pre- and postnatal nicotine treatment did not affect the overall sexual performance of the male rats, although the number of mounts and intromissions during a second series decreased. Eye opening of male and female pups was accelerated with pre/postnatal nicotine administration. Birth weight, testis and levator ani weights, sex ratio and number of pups per litter were not affected. We suggest that nicotine may act as a neuromodulator during sexual differentiation of the brain, demasculinizing the male progeny in rats. This effect is correlated with decreased testosterone levels during adulthood.

Tamoxifen and estradiol improved locomotor function and increased spared tissue in rats after spinal cord injury: Their antioxidant effect and role of estrogen receptor alpha

17β-Estradiol is a multi-active steroid that imparts neuroprotection via diverse mechanisms of action. However, its role as a neuroprotective agent after spinal cord injury (SCI), or the involvement of the estrogen receptor-alpha (ER-α) in locomotor recovery, is still a subject of much debate. In this study, we evaluated the effects of estradiol and of Tamoxifen (an estrogen receptor mixed agonist/antagonist) on locomotor recovery following SCI. To control estradiol cyclical variability, ovariectomized female rats received empty or estradiol filled implants, prior to a moderate contusion to the spinal cord. Estradiol improved locomotor function at 7, 14, 21, and 28 days post injury (DPI), when compared to control groups (measured with the BBB open field test). This effect was ER-α mediated, because functional recovery was blocked with an ER-α antagonist. We also observed that ER-α was up-regulated after SCI. Long-term treatment (28 DPI) with estradiol and Tamoxifen reduced the extent of the lesion cavity, an effect also mediated by ER-α. The antioxidant effects of estradiol were seen acutely at 2 DPI but not at 28 DPI, and this acute effect was not receptor mediated. Rats treated with Tamoxifen recovered some locomotor activity at 21 and 28 DPI, which could be related to the antioxidant protection seen at these time points. These results show that estradiol improves functional outcome, and these protective effects are mediated by the ER-α dependent and independent-mechanisms. Tamoxifen׳s effects during late stages of SCI support the use of this drug as a long-term alternative treatment for this condition.

Estrogen Influences Cocaine-Induced Blood Oxygen Level-Dependent Signal Changes in Female Rats

We investigated the effect of estrogen on cocaine-induced brain activity using blood oxygen level-dependent (BOLD) magnetic resonance imaging. Ovariectomized (Ovx) rats without estrogen and Ovx rats with estrogen (Ovx+E) were given a single saline or cocaine injection (15 mg/kg, i.p.) for 5 d. After 7 d of withdrawal from injections, rats were challenged with cocaine during functional imaging. Acute cocaine administration produced positive BOLD activation in the prefrontal cortex, nucleus accumbens, striatum, ventral tegmental area, and hippocampus, among other brain regions. Positive BOLD signal changes were lower in Ovx+E than in Ovx rats. With repeated cocaine administration, Ovx+E rats showed enhanced BOLD signal changes in the nucleus accumbens, ventral tegmental area, and hippocampus compared with acutely treated animals. Our results indicate that estrogen influences the effects of acute and repeated cocaine administration on BOLD signal changes. The data suggest that in females with estrogen, cocaine-induced neuronal activity is enhanced after repeated cocaine administration. It is possible that the actions of estrogen within the aforementioned brain regions potentiate the behavioral response to cocaine observed in female rats.