José R. Criado

Mice devoid of prion protein have cognitive deficits that are rescued by reconstitution of PrP in neurons

Prion protein (PrP(C)) is a constituent of most normal mammalian cells and plays an essential role in the pathogenesis of transmissible spongiform encephalopathies (TSE). However, the normal cellular function of PrP(C) remains unclear. Here, we document that mice with a selective deletion of PrP(C) exhibited deficits in hippocampal-dependent spatial learning, but non-spatial learning remained intact. mPrP-/- mice also showed reduction in paired-pulse facilitation and long-term potentiation in the dentate gyrus in vivo. These deficits were rescued in transgenic mPrP-/- mice expressing PrP(C) in neurons under control of the neuron-specific enolase (NSE) promoter indicating that they were due to lack of PrP(C) function in neurons. The deficits were seen in mPrP-/- mice with a homogeneous 129/Ola background and in mPrP-/- mice in the mixed (129/Ola x C57BL/10) background indicating that these abnormalities were unlikely due to variability of background genes or alteration of the nearby Prnd (doppel) gene.

Differential sensitivity of c-Fos expression in hippocampus and other brain regions to moderate and low doses of alcohol.

Alcohol consumption in humans is characterized by a wide range of behavioral effects and pathological consequences that suggest several neuroanatomical targets for this drug. To identify these targets we have mapped alcohol-induced changes in the expression of the c-Fos protein in the rat brain. Administration of a moderate dose of alcohol (1.5 g kg−1) led to a suppression of basal and novel environment-induced c-Fos expression in the hippocampus and simultaneous induction of this protein in regions important for the reinforcing as well as aversive properties of drugs. These include the extended amygdala (including the central nucleus of amygdala, bed nucleus of stria terminalis and nucleus accumbens), regions processing sensory information (including the Edinger–Westphal nucleus and the paraventricular nucleus of the thalamus) and in stress-related areas (including the paraventricular nucleus of the hypothalamus, nucleus of the solitary tract and several neocortical areas). Repeated administration of the same dose of alcohol did not decrease alcohol-mediated suppression of c-Fos in the hippocampus, but decreased alcohol-induced expression of c-Fos in other areas. A lower dose of acute alcohol (0.5 g kg−1) reduced basal c-Fos expression in several areas of the neocortex, hippocampus and hypothalamus. However, while this low dose of alcohol was unable to counteract the environmental novelty-induced c-Fos expression in these areas, it increased c-Fos expression in the central nucleus of amygdala (an effect similar to the one observed previously for diazepam). Our data suggest that the effects of low doses of alcohol may be due to selective GABA-like effects of ethanol, whereas higher doses of ethanol involve effects on multiple neurotransmitter systems.

Profound increase in sensitivity to glutamatergic- but not cholinergic agonist-induced seizures in transgenic mice with astrocyte production of IL-6.

Transgenic mice with glial fibrillary acidic protein (GFAP) promoter driven‐astrocyte production of the cytokines interleukin‐6 (IL‐6) and tumor necrosis factor (TNF) were used to determine whether the pre‐existing production of these cytokines in vivo might modulate the sensitivity of neurons to excitotoxic agents. Low doses of kainic acid (5 mg/kg) that produced little or no behavioral or electroencephalogram (EEG) alterations in wild type or glial fibrillary acidic protein (GFAP)‐TNF animals induced severe tonic‐clonic seizures and death in GFAP‐IL6 transgenic mice of 2 or 6 months of age. GFAP‐IL6 mice were also significantly more sensitive to N‐methyl‐D‐aspartate (NMDA)‐ but not pilocarpine‐induced seizures. Kainic acid uptake in the brain of the GFAP‐IL6 mice was higher in the cerebellum but not in other regions. Kainic acid binding in the brain of GFAP‐IL6 mice had a similar distribution and density as wild type controls. In the hippocampus of GFAP‐IL6 mice that survived low dose kainic acid, there was no change in the extent of either neurodegeneration or astrocytosis. Immunostaining revealed degenerative changes in gamma aminobutyric acid (GABA)‐ and parvalbumin‐positive neurons in the hippocampus of 2‐month‐old GFAP‐IL6 mice which progressed to the loss of these cells at 6 months of age. Thus, GFAP‐IL6 but not GFAP‐TNF mice showed markedly enhanced sensitivity to glutamatergic‐ but not cholinergic‐induced seizures and lethality. This may relate, in part, to a compromise of inhibitory interneuron function. Therefore, pre‐existing IL‐6 production and inflammation in the central nervous system (CNS) not only causes spontaneous neurodegeneration but also synergizes with other neurotoxic insults to induce more severe acute functional neurological impairment.

Age-independent and age-related deficits in visuospatial learning, sleep-wake states, thermoregulation and motor activity in PDAPP mice.

Recent studies demonstrated that mice overexpressing the human mutant beta-amyloid precursor protein (hbetaAPP; PDAPP mice) show age-independent and age-related deficits in spatial learning. We used behavioral and electrophysiological techniques to determine in young and aged PDAPP mice whether deficits in spatial learning also involve alterations in sleep-wake states, thermoregulation and motor activity. Consistent with earlier studies, young PDAPP mice exhibited selective age-independent deficits using spatial, but not random and serial strategies in the circular maze. Aged PDAPP mice exhibited deficits using all search strategies. The core body temperature (Tb) in young and aged PDAPP mice was significantly lower than in age-matched non-transgenic (non-Tg) littermates. During the dark period, the motor activity (LMA) was significantly increased in young PDAPP mice, but not in aged PDAPP mice. During the light period, young PDAPP mice showed a reduction in the generation of rapid-eye-movement (REM) sleep. In contrast, aged PDAPP mice exhibited a reduction in the amount of time spent in W and an increase in SWS during the light period. Aged PDAPP mice also showed an increase in the amount of time spent in W and a reduction in REM sleep during the dark period. Our findings support previous reports indicating deficits in spatial learning in young and aged PDAPP mice. These data also suggest that PDAPP mice exhibit age-independent and age-related deficits in neural mechanisms regulating visuospatial learning, the total amount and the circadian distribution of sleep-wake states, thermoregulation and motor activity.

In vivo synaptic transmission in young and aged amyloid precursor protein transgenic mice.

Alzheimers disease (AD) is characterized by progressive neurodegeneration and cognitive impairment. We examined in vivo alterations in hippocampal neurotransmission in both young and aged PDAPP transgenic mice and nontransgenic littermates. We now report that in vivo abnormal neurotransmission in hippocampal circuits of PDAPP mice precedes beta deposition and neurodegeneration. These in vivo data provide the first evidence that dysfunction in hippocampal neuronal circuits may not be correlated with age-related extracellular beta plaque deposition.

Urotensin II Modulates Rapid Eye Movement Sleep through Activation of Brainstem Cholinergic Neurons

Urotensin II (UII) is a cyclic neuropeptide with strong vasoconstrictive activity in the peripheral vasculature. UII receptor mRNA is also expressed in the CNS, in particular in cholinergic neurons located in the mesopontine tegmental area, including the pedunculopontine tegmental (PPT) and lateral dorsal tegmental nuclei. This distribution suggests that the UII system is involved in functions regulated by acetylcholine, such as the sleep-wake cycle. Here, we tested the hypothesis that UII influences cholinergic PPT neuron activity and alters rapid eye movement (REM) sleep patterns in rats. Local administration of UII into the PPT nucleus increases REM sleep without inducing changes in the cortical blood flow. Intracerebroventricular injection of UII enhances both REM sleep and wakefulness and reduces slow-wave sleep 2. Intracerebroventricular, but not local, administration of UII increases cortical blood flow. Moreover, whole-cell recordings from rat-brain slices show that UII selectively excites cholinergic PPT neurons via an inward current and membrane depolarization that were accompanied by membrane conductance decreases. This effect does not depend on action potential generation or fast synaptic transmission because it persisted in the presence of TTX and antagonists of ionotropic glutamate, GABA, and glycine receptors. Collectively, these results suggest that UII plays a role in the regulation of REM sleep independently of its cerebrovascular actions by directly activating cholinergic brainstem neurons.

Acute ethanol induces c-fos immunoreactivity in GABAergic neurons of the central nucleus of the amygdala

The central nucleus of the amygdala (CNA) is a component of the brain reward pathway which is believed to represent an anatomical substrate for drugs of abuse. Previous studies have shown that acute ethanol administration induces the expression of c-fos in the CNA of rat brains. We report here, that over 70% of these c-fos immunoreactive neurons are GABAergic. This observation provides the first anatomical evidence that GABAergic neurons of the CNA are responsive to acute ethanol exposure and suggest that the GABAergic system of the CNA is a key neuronal substrate for ethanol actions on the central nervous system.

Adolescent ethanol exposure: does it produce long-lasting electrophysiological effects?

This review discusses evidence for long-lasting neurophysiological changes that may occur following exposure to ethanol during adolescent development in animal models. Adolescence is the time that most individuals first experience ethanol exposure, and binge drinking is not uncommon during adolescence. If alcohol exposure is neurotoxic to the developing brain during adolescence, not unlike it is during fetal development, then understanding how ethanol affects the developing adolescent brain becomes a major public health issue. Adolescence is a critical time period when cognitive, emotional, and social maturation occurs and it is likely that ethanol exposure may affect these complex processes. To study the effects of ethanol on adolescent brain, animal models where the dose and time of exposure can be carefully controlled that closely mimic the human condition are needed. The studies reviewed provide evidence that demonstrates that relatively brief exposure to high levels of ethanol, via ethanol vapors, during a period corresponding to parts of adolescence in the rat is sufficient to cause long-lasting changes in functional brain activity. Disturbances in waking electroencephalogram and a reduction in the P3 component of the event-related potential (ERP) have been demonstrated in adult rats that were exposed to ethanol vapor during adolescence. Adolescent ethanol exposure was also found to produce long-lasting reductions in the mean duration of slow-wave sleep (SWS) episodes and the total amount of time spent in SWS, a finding consistent with a premature aging of sleep. Further studies are necessary to confirm these findings, in a range of strains, and to link those findings to the neuroanatomical and neurochemical mechanisms potentially underlying the lasting effects of adolescent ethanol exposure.

Acute ethanol induction of c-Fos immunoreactivity in pre-pro-enkephalin expressing neurons of the central nucleus of the amygdala

Previous studies have shown that acute ethanol administration induces expression of c-Fos immunoreactivity in the central nucleus of the amygdala (CNA) [S.L. Chang, N.A. Patel, A.A. Romero, Activation and desensitization of Fos immunoreactivity in the rat brain following ethanol administration, Brain Res., 679 (1995) 89-98; M. Morales, J.R. Criado, P.P. Sanna, S.J. Henriksen, F.E. Bloom, Acute ethanol induces c-fos immunoreactivity in GABAergic neurons of the central nucleus of the amygdala, Brain Res., 798 (1998) 333-336; A.E. Ryabinin, J.R. Criado, S.J. Henriksen, F.E. Bloom, M.C. Wilson, Differential sensitivity of c-Fos expression in hippocampus and other brain regions to moderate and low doses of alcohol, Mol. Psychiatry, 2 (1997) 32-43]. We recently showed that over 70% of these c-Fos immunoreactive neurons are GABAergic [M. Morales, J.R. Criado, P.P. Sanna, S.J. Henriksen, F.E. Bloom, Acute ethanol induces c-fos immunoreactivity in GABAergic neurons of the central nucleus of the amygdala, Brain Res, 798 (1998) 333-336]. In the present study, we report that ethanol-induced c-Fos immunoreactivity was mainly confined to neurons that express pro-enkephalin (ENK). In contrast, a small number of c-Fos immunoreactive neurons express corticotrophin releasing factor (CRF). Our results thus provide anatomical evidence indicating that within the amygdala, GABAergic neurons that contain ENK are responsive to acute ethanol exposure.

Structural and Compositional Determinants of Cortistatin Activity

Cortistatin‐14 (CST‐14) is a putative novel neuropeptide that shares 11 of its 14 residues with somatostatin‐14 (SRIF‐14), yet its effects on sleep physiology, locomotor behavior and hippocampal function are different from those of somatostatin. We studied the structural basis for cortistatins distinct biological activities. As with SRIF‐14, CST‐14 does not show any preferred conformation in solution, as determined by circular dichroism and nuclear magnetic resonance. Synthetic cortistatin analogs were designed and synthesized based on the cyclic structure of octreotide. Biological assays were carried out to determine their binding affinities to five somatostatin receptors (sst1‐5) and their ability to produce changes in locomotor activity and to modulate hippocampal physiology and sleep. The results show that the compound with N‐terminal proline and C‐terminal lysine amide exhibits cortistatin‐like biological activities, including reduction of population spike amplitudes in the hippocampal CA1 region, decrease in locomotor activity and enhancement of slow‐wave sleep 2. These findings suggest that both proline and lysine are necessary for cortistatin binding to its specific receptor. J. Neurosci. Res. 56:611–619, 1999.