Prestin autoantibodies screening in idiopathic sudden sensorineural hearing loss.

OBJECTIVES: To define the clinical association of serum prestin autoantibodies and their impact on prognosis, as specific serum diagnostic markers in patients with idiopathic sudden sensorineural hearing loss (ISSNHL). DESIGN: Sera from 63 patients with ISSNHL were screened prospectively for the presence of prestin autoantibodies by an enzyme-linked immunosorbent assay (Elisa) test. Serum was assayed for anti-prestin IgG antibodies using recombinant human prestin (SLC26 A5). Demographic, clinical, and audiometric variables were analyzed. RESULTS: Two patients (3.17%) had demonstrable anti-prestin antibodies in serum (exact 95% CI: -1.16% to 7.5%). No statistically significant association was found between prestin autoantibodies and demographic or audiologic parameters. CONCLUSIONS: This preliminary and novel study does not support the presence of an active humoral immune reaction against prestin in ISSNHL.

The function of the adrenocortical axis in permanent middle cerebral artery occlusion: effect of glucocorticoids on the neurological outcome.

We characterized the effect of acute ischemic stroke on the activation of the hypothalamic-pituitary-adrenal (HPA) axis and evaluated the role of glucocorticoids (GC) in the clinical outcome following ischemic stroke. Male spontaneous hypertensive rats underwent permanent middle cerebral artery occlusion (PMCAO) and developed a cortical infarct. At 4h post-PMCAO or sham operation, serum levels of ACTH and corticosterone (CS) were elevated 5 and 4 fold respectively as compared to controls and then returned to basal levels at 24h post surgery. In these experimental groups we found also a significant depletion of median eminence (ME)-CRH(41). In adrenalectomized (Adx) rats that underwent PMCAO the degree of motor disability and infarct volume was similar to that of intact rats. Administration of dexamethasone (Dex) to Adx-PMCAO rats significantly improved the motor disability and decreased the infarct volume. However, in sham-Adx with PMCAO, Dex had no effect on these two parameters. In rats with PMCAO or sham-PMCAO, brain production of PGE(2) was significantly increased. This effect was further enhanced in Adx-PMCAO rats and significantly inhibited by Dex. In conclusion, activation of the HPA axis following PMCAO is due to stress induced by surgery. This activation is mediated by hypothalamic CRH(41). Absence of endogenous GC or administration of Dex in naïve rats does not alter motor and pathological parameters in the acute stage following PMCAO. In contrast, administration of Dex significantly improved the outcome following cerebral ischemia in Adx rats which may be due to increased glucocorticoid receptors. Brain production of PGE(2) does not play an important role in the pathophysiology of the acute phase of cerebral ischemia.

Copaxone interferes with the PrP Sc-GAG interaction.

The hallmark of prion disease-induced neurodegeneration is the accumulation of PrP(Sc), a misfolded form of PrP(C). In addition, several lines of evidence indicate a role for the immune system and, in particular, inflammation in prion disease pathogenesis. In this work, we tested whether Copaxone, an immunomodulatory agent currently used for the treatment of multiple sclerosis, can affect prion disease manifestation in scrapie-infected hamsters. We show here that Copaxone exerted no effect on prion disease incubation time when treatment commenced 2 weeks after i.p. prion infection. However, when Copaxone was mixed with the initial prion inoculum or administered to hamsters weekly starting on the day of infection, prion disease incubation time was prolonged by 30 days. This suggests that Copaxone may affect the initial infection process. In vitro experiments indicate that Copaxone significantly reduced PrP(Sc) binding to both Chinese hamster ovary (CHO) cells and heparin beads and also binds to heparin by itself. Interestingly, Copaxone also abolished PrP(Sc) accumulation in scrapie-infected cells. We propose that Copaxone delays prion infection by competing with the PrP(Sc)-glycosaminoglycans interaction. Whether the immunomodulating activity of Copaxone is related to its heparin binding and anti-prion properties remains to be established.

Rasmussen encephalitis with active inflammation and delayed seizures onset.

Severe focal epilepsy is regarded as a clinical hallmark of Rasmussen encephalitis (RE). The authors report two children with progressive hemiparesis, contralateral hemispheric atrophy, and pathologic features characteristic for RE. At histologic diagnosis and over several months, neither patient experienced seizures. The report enlarges the clinical spectrum of RE and suggests that seizures are not an obligatory presenting symptom of the disorder.

Infantile bilateral striatal necrosis maps to chromosome 19q.

BACKGROUND: Infantile bilateral striatal necrosis (IBSN) encompasses several syndromes of bilateral symmetric degeneration of the caudate nucleus, putamen, and globus pallidus. Autosomal recessive IBSN is characterized clinically by developmental arrest beginning at age 7 to 15 months, dysphagia, choreoathetosis, pendular nystagmus and optic atrophy, and severe progressive atrophy of the basal ganglia on MRI. OBJECTIVE: To map the gene causing IBSN. METHODS: A 10-cM genome-wide linkage scan was initially performed on five affected and five unaffected individuals. The extended family was included in the analysis to narrow the candidate region. Logarithm of odds (LOD) score was calculated using the SUPERLINK program. RESULTS: Linkage to the chromosomal region 19q13.32-13.41 was established (Z(max) = 6.27 at theta = 0.02 at locus D19S412). Recombination events and a common disease-bearing haplotype defined a critical region of 1.2 Mb between the loci D19S596 proximally and D19S867 distally. CONCLUSION: IBSN maps to the chromosomal region 19q13.32-13.41. The presence of a common haplotype in all the patients suggests that the disease is caused by a single mutation derived from a single ancestral founder in all the families.

Combination of dexanabinol and tempol in focal cerebral ischemia: is there a ceiling effect?

Because ischemic neuronal death is triggered by several parallel mechanisms, a combination of drugs active against individual death-promoting mechanisms may have synergistic effects. Dexanabinol is a noncompetitive NMDA antagonist with anti-inflammatory effects and tempol is a nitroxide antioxidant. Therefore, we explored whether their combined use results in smaller infarct volumes as compared with their individual administration. Rats underwent permanent middle cerebral artery occlusion (PMCAO) and were given vehicle, dexanabinol alone, tempol alone, or a combination of dexanabinol and tempol (n = 13 per group) 1 h later. Five animals in each group were evaluated with a motor rating scale 24 h after PMCAO and the infarct volumes were then measured. The remaining animals were examined with motor and behavioral scales up to 30 days after PMCAO and their infarct volumes were then determined. Motor disability and water maze latencies at all time points examined and infarct volumes at days 1 and 30 were significantly reduced in all active treatment groups when compared with vehicle. However, no significant differences were observed between the active treatment groups. In conclusions, combination therapy with dexanabinol and tempol does not appear to have additional neuroprotective effects compared to those conferred by each agent alone even when administered at optimal timing and dosing. Therefore, a ceiling neuroprotective effect that is impossible to overcome may exist.

The EAE-associated behavioral syndrome: I. Temporal correlation with inflammatory mediators.

To elucidate the mechanisms underlying the EAE-associated behavioral syndrome (EBS), we examined the temporal correlation between the behavioral alterations and inflammatory processes. Onset of the behavioral syndrome was associated with the onset of brain infiltration, as well as mRNA expression of interleukin 1 beta (IL-1 beta) and tumor necrosis factor alpha (TNF-alpha) and elevated production of interleukin 1 beta protein and prostaglandin E(2) (PGE(2)). Sickness behavior symptoms coincided with peak cytokine expression. Behavioral recovery was associated with a reduction of cytokine expression, but not infiltration, PGE(2) production or motor disturbances. These results suggest that inflammatory processes in general, and the production of pro-inflammatory cytokines in particular, are involved in mediating EAE-associated sickness behavior.

The EAE-associated behavioral syndrome: II. Modulation by anti-inflammatory treatments.

EAE is associated with sickness behavior symptoms that are temporally correlated with inflammatory processes. To further elucidate the role of inflammatory mediators in the behavioral syndrome, EAE mice were injected daily with anti-inflammatory drugs, beginning at disease onset. Dexamethasone or interleukin-1 (IL-1) receptor antagonist or the prostaglandins synthesis inhibitor indomethacin attenuated the behavioral symptoms. Administration of the tumor necrosis-factor alpha (TNF-alpha) synthesis inhibitor pentoxifylline or targeted deletion of the type I TNF receptor had no behavioral effects whereas administration of pentoxifylline in IL-1ra-treated mice further reversed the behavioral depression. These findings demonstrate the critical involvement of pro-inflammatory cytokines and prostaglandins in the EAE-associated behavioral syndrome, and may have implications for understanding and treating the neuropsychiatric disturbances in multiple sclerosis (MS) patients.

Experimental autoimmune encephalomyelitis-associated behavioral syndrome as a model of 'depression due to multiple sclerosis'.

Many medical conditions, including inflammatory diseases such as multiple sclerosis (MS), are often accompanied by a high prevalence of depressive episodes. Inflammatory mediators, such as cytokines, were implicated in illness-associated depressive conditions, both in humans and in animals. For example, MS-associated depression (MSD) was attributed to pathophysiological processes such as immune dysregulation and cerebral inflammation. We have recently documented a depressive-like behavioral syndrome in mice with experimental autoimmune encephalomyelitis (EAE), an established model of MS. In the present paper, we discuss the similarities between the EAE-associated behavioral syndrome (EBS) and MSD, in terms of phenomenology, putative mechanisms and responsiveness to anti-depressive therapy. In particular, we show that: (1) EAE and depression are associated not only with similar behavioral symptomatology, but also with common physiological alterations, including impaired serotonergic neurotransmission, and activation of neuroendocrine (e.g., adrenocortical) and inflammatory cytokine systems; (2) the EBS precedes any neurological deficit during the initial EAE attack, as well as further exacerbations, and remits during recovery and between relapses. Similarly, in many MS patients depression precedes and accompanies the attacks and wanes during remissions; (3) females show increased susceptibility to EBS. Similarly, depression is much more prevalent in women than in men; (4) chronic treatment with the tricyclic anti-depressant imipramine reduced EAE-induced mortality, body-weight loss and behavioral suppression. Similarly, anti-depressant drugs have been used effectively in treating MSD. These findings suggest that the EBS may serve as an animal model for MSD.

The nitroxide antioxidant tempol is cerebroprotective against focal cerebral ischemia in spontaneously hypertensive rats.

Free radicals appear to participate in the final common pathway of neuronal death in ischemia and may therefore be an adequate target for therapy. Tempol is a nitroxide antioxidant with proven protective efficacy in several animal models, including myocardial ischemia, that has not been previously tested in models of permanent cerebral ischemia. Spontaneously hypertensive rats underwent permanent middle cerebral artery occlusion (PMCAO). Following dose-response and time-window-finding experiments rats were given vehicle or tempol (50 mg/kg) subcutaneously 1 h after PMCAO (n = 10/group). Five animals in each group were evaluated with a motor scale 24 h after the infarct and were then sacrificed and the injury volume was measured. The remaining animals were examined daily with the motor scale and also with a Morris water maze test on days 26-30 after PMCAO and sacrificed on day 30. Motor scores at all time points examined were significantly better in the tempol-treated animals (P < 0.05 for all). Significantly better performance in the water maze test for performance on days 26-30 was noted in the tempol group compared with the vehicle-treated group (P < 0.05). Injury volumes at days 1 and 30 were significantly reduced in the tempol group (9.83 +/- 1.05 vs 19.94 +/- 1.43% hemispheric volume, P = 0.0009, and 13.2 +/- 2.97 vs 24.4 +/- 2.38% hemispheric volume, P = 0.02, respectively). In conclusion, treatment with tempol led to significant motor and behavioral improvement and reduced injured tissue volumes both in the short and in the long term after stroke.

Expression of endothelial nitric oxide synthase in the ischemic penumbra: relationship to expression of neuronal nitric oxide synthase and vascular endothelial growth factor.

Expressional patterns of the endothelial and neuronal forms of nitric oxide synthase (NOS) in cerebral ischemia were studied utilizing a permanent middle cerebral artery occlusion (PMCAO) model. Motor performance and infarct volumes were determined in the rats. Immunohistochemical staining for eNOS, nNOS and neurofilament were performed at 1, 2, 3, 5, 7 and 14 days after PMCAO. Vascular endothelial growth factor (VEGF) expression was determined by in-situ hybridization. PMCAO caused a reproducible cortical infarct with motor deficits in the rats. Double immunohistochemical stainings indicated that eNOS and nNOS were induced in ischemic neurons. Most stained neurons were positive for both NOS forms but some reacted with only one NOS antibody. nNOS expression peaked at 24-48 h after PMCAO, stained mainly the cytoplasm of core neurons, and disappeared after the 3rd day. eNOS expression increased until the 7th day, stained mainly the cytoplasm and membrane of penumbral cells and disappeared by the 14th day after PMCAO. VEGF expression was significantly induced in the penumbral zone in a similar distribution to eNOS. The anatomical and temporal pattern of VEGF and eNOS induction in the brain after permanent ischemia suggest that these mediators may play a role in protecting penumbral tissue from additional ischemic damage.

Evaluation of the effect of stress on the blood--brain barrier: critical role of the brain perfusion time.

The Gulf war syndrome has drawn increased attention in the issue of the effect of stress on the blood-brain barrier (BBB). We have applied various stressful modalities and tested BBB disruption as measured by the amount of Evans blue (EB) retained by brain parenchyma. We have evaluated the retention of this marker as a function of the perfusion time of the brain following stress. This was done to distinguish between the marker retained in the lumen of small blood vessels and the marker retained by the brain parenchyma. Mice were exposed to either short swim stress or restraint stress. In mice exposed to either swim or restraint stress that were perfused for 1 min, the amount of EB retained in the brain was significantly higher as compared to non-stressed controls. Fifteen min perfusion markedly reduced the EB brain content to levels found in the non-stressed animals. In rats exposed to neural or metabolic stressful stimuli and perfused for 15 min, the EB content was similar to non-stressed controls. Our results demonstrate that various stress modalities have no effect on the BBB permeability and insufficient wash of blood vessels by perfusion may cause misinterpretation of permeability studies.

Long term cerebroprotective effects of dexanabinol in a model of focal cerebral ischemia.

In order to test the long-term cerebroprotective effects of dexanabinol, a synthetic non-competitive NMDA antagonist that also has anti-TNFalpha effects, spontaneously hypertensive rats underwent permanent middle cerebral artery occlusion (PMCAO). Rats were given vehicle or dexanabinol (4.5 mg/kg) 1, 3 or 6 h after PMCAO. The research consisted of 2 stages. In the short-term set of experiments animals (n=5/group), were tested with a motor disability scale 24 h post PMCAO, then sacrificed and the infarct volume was measured using 2,3,5-Triphenyltetrazolium chloride (TTC) staining. In the long-term set of experiments the rats (n=7/group) were examined daily with a motor disability scale up to 30 days after PMCAO and then sacrificed and infarct volumes were determined using TTC staining. Motor scores were significantly improved in the dexanabinol treated rats (P<0.05 for all groups) at all the time points examined. Infarct volumes were significantly reduced 24 h after PMCAO in the groups treated 1 or 3 h, but not 6 h after PMCAO compared with vehicle (Mean+/-S.D., 11.5+/-2.02, 12+/-3.2 and 14.4+/-2.4% vs. 20.8+/-1.3% hemispheric volume respectively). The lesions remained significantly smaller in the dexanabinol groups 30 days after PMCAO (Mean+/-S.D., 24.49+/-1.9% vs. 8.1+/-0.6, 11.1+/-2.3 and 13.8+/-2.5% hemispheric volume in animals treated with vehicle vs. dexanabinol 1, 3 or 6 h after PMCAO respectively; P<0.05 for all). In conclusion, the extended therapeutic window and the multi-mechanistic durable neuroprotective effects of dexanabinol make it a promising candidate for future stroke therapy.

Effects of antidepressant drugs on the behavioral and physiological responses to lipopolysaccharide (LPS) in rodents.

Antidepressants produce various immunomodulatory effects, as well as an attenuation of the behavioral responses to immune challenges, such as lipopolysaccharide (LPS). To explore further the effects of antidepressants on neuroimmune interactions, rats were treated daily with either fluoxetine (Prozac) or saline for 5 weeks, and various behavioral, neuroendocrine, and immune functions were measured following administration of either LPS or saline. Chronic fluoxetine treatment significantly attenuated the anorexia and body weight loss, as well as the depletion of CRH-41 from the median eminence and the elevation in serum corticosterone levels induced by LPS. Chronic treatment with imipramine also attenuated LPS-induced adrenocortical activation. In rats and in mice, which normally display a biphasic body temperature response to LPS (initial hypothermia followed by hyperthermia), chronic treatment with fluoxetine completely abolished the hypothermic response and facilitated and strengthened the hyperthermic response. The effects of antidepressants on the responsiveness to LPS are probably not mediated by their effects on peripheral proinflammatory cytokine production, because LPS-induced expression of TNFalpha and IL-1beta mRNA in the spleen (assessed by semiquantitative in situ hybridization) was not altered following chronic treatment with either fluoxetine or imipramine. The effects of antidepressants on the acute phase response may have important clinical implications for the psychiatric and neuroendocrine disturbances that are commonly associated with various medical conditions.

Renal nitric oxide production during the early phase of experimental diabetes mellitus.

BACKGROUND: Diabetic nephropathy (DN) is characterized by hyperfiltration and hypertrophy in experimental models of diabetes mellitus (DM). Several studies have demonstrated that the pathophysiologic and morphologic changes in DN are mediated by either an increase or decrease in renal nitric oxide (NO) production and/or activity. The goal of the present study was to determine the effects that the early diabetic state has on NO production in the kidney of rats with streptozotocin-induced DM. METHODS: Experimental DM was induced in rats with streptozotocin. Urinary NO production was measured, and levels and activity of the different NOS isoforms were determined by a combination of techniques, including immunoblotting, immunohistochemistry, diaphorase staining, and reverse transcription-polymerase chain reaction. RESULTS: During the first week of DM, urinary NO metabolites (uNO2 + NO3) were reduced as compared with controls, which were unrelated to changes in serum levels of NO. Total NO synthase (NOS) activity was reduced in the renal cortex beginning at 30 hours after the induction of DM. NADPH diaphorase staining of renal cortical slices showed reduced NOS activity in the macula densa in diabetic animals. By immunohistochemical staining with antibodies to the different isoforms of NOS, it was found that protein levels of the neuroneal NOS (nNOS) isoform was diminished in the macula densa. No changes were found in the levels of endothelial NOS (eNOS) activity and protein in the renal cortex in the early diabetic state. CONCLUSIONS: This study provides strong evidence that renal production of NO is reduced in early DM and that this reduction is associated with decreased levels of nNOS activity and protein in the macula densa.

Behavioral aspects of experimental autoimmune encephalomyelitis.

Acute inflammation is known to induce a depressive-like sickness behavior syndrome in humans and in experimental animals. In the present study, we sought to determine whether a chronic neuroautoimmune inflammation is also associated with a similar behavioral syndrome. Experimental autoimmune encephalomyelitis (EAE) was induced in SJL/J female mice by adoptive transfer of lymph node cells, and sickness behavior symptoms, including anorexia, loss of body weight, reduced social exploration, and decreased preference for sucrose solution were measured. We report that these components of sickness behavior were induced during the acute phase of the disease, and recovered in later phases. Moreover, the onset and recovery of the behavioral symptoms preceded the onset and recovery of the neurological signs, respectively. Since EAE is considered a model for multiple sclerosis (MS), it is suggested that EAF-induced behavioral changes may serve as a model for the depressive symptomatology that characterizes most MS patients.

Illness, cytokines, and depression.

Various medical conditions that involve activation of the immune system are associated with psychological and neuroendocrine changes that resemble the characteristics of depression. In this review we present our recent studies, designed to investigate the relationship between the behavioral effects of immune activation and depressive symptomatology. In the first set of experiments, we used a double-blind prospective design to investigate the psychological consequences of illness in two models: (1) vaccination of teenage girls with live attenuated rubella virus, and (2) lipopolysaccharide (LPS) administration in healthy male volunteers. In the rubella study, we demonstrated that, compared to control group subjects and to their own baseline, a subgroup of vulnerable individuals (girls from low socioeconomic status) showed a significant virus-induced increase in depressed mood up to 10 weeks after vaccination. In an ongoing study on the effects of LPS, we demonstrated significant LPS-induced elevation in the levels of depression and anxiety as well as memory deficits. These psychological effects were highly correlated with the levels of LPS-induced cytokine secretion. In parallel experiments, we demonstrated in rodents that immune activation with various acute and chronic immune challenges induces a depressive-like syndrome, characterized by anhedonia, anorexia, body weight loss, and reduced locomotor, exploratory, and social behavior. Chronic treatment with antidepressants (imipramine or fluoxetine) attenuated many of the behavioral effects of LPS, as well as LPS-induced changes in body temperature, adrenocortical activation, hypothalamic serotonin release, and the expression of splenic TNF-alpha mRNA. Taken together, these findings suggest that cytokines are involved in the etiology and symptomatology of illness-associated depression.

Effect of exogenous nitric oxide and inhibitors of nitric oxide synthase on the hypothalamic pituitary adrenal axis responses to neural stimuli.

It has been shown that the hypothalamic-pituitary-adrenal (HPA) axis responses to immune-derived stimuli in particular can be modulated by nitric oxide (NO). In the present study we examined the effect of endogenous and exogenous NO on the HPA axis responses to neural stimuli which are not related to immune functions. Intracerebroventricular injection of NOR-3, a donor of NO, had no effect on basal HPA axis activity but significantly attenuated the secretion of median eminence (ME) CRH-41 as well as the serum ACTH and corticosterone (CS) in response to acute photic stimulation in a dose-dependent manner. Intracerebroventricular administration of N-omega-nitro-L-arginine methyl ester (L-NAME), a general NOS inhibitor, significantly enhanced ACTH and CS responses to this stress but did not change the basal levels of these hormones. On the other hand, i.c.v. injection of aminoguanidine, an inhibitor of inducible NO synthase (NOS) but not of neuronal NOS, did not affect the HPA axis responses to photic stimulation. These results suggest that: (1) NO is involved in modulation of the HPA axis responses to neural stimuli which are not dependent on immune factors, (2) the effect of NO is mediated by inhibition of hypothalamic ME CRH-41 secretion, and (3) this effect is probably mediated by neuronal NOS and not by inducible NOS.