Young adult ischaemic stroke related acute symptomatic and late seizures: risk factors.

BACKGROUND AND PURPOSE: After first-ever ischaemic stroke, to assess the risk and baseline factors associated with acute symptomatic seizure (ASS) (occurring within 7 days) and late post-stroke seizure (LPS) (>7 days). METHODS: All consecutive patients aged 15-49 with first-ever ischaemic stroke between 1994 and 2007 treated at the Helsinki University Central Hospital were included, using Cox proportional hazard models to identify factors associated with seizures. Adjustment was for age, gender, vascular risk factors, admission hyperglycemia (>6.1 mm) and hyponatremia (<137 mm), use of psychiatric medication, stroke severity (NIH Stroke Scale) and anatomical (Bamford criteria) and etiological (Trial of Org in Acute Stroke Treatment) stroke subtype. RESULTS: ASSs emerged in 35 (3.5%) patients. LPSs (n = 102) occurred at a cumulative rate of 6.1% at 1 year, 9.5% at 5 years and 11.5% at 10 years. In multivariate analysis, anxiolytic use at time of index stroke (hazard ratio 13.43, 95% confidence interval 3.91-46.14), moderate stroke severity (3.95, 1.86-8.41), cortical involvement (3.69, 1.66-8.18) and hyponatremia (3.26, 1.41-7.57) were independently associated with ASSs. Risk factors for LPSs were total anterior circulation infarct (15.94, 7.62-33.33), partial anterior circulation infarct (3.48, 1.52-7.93), history of ASS (3.94, 2.07-7.48), antidepressant use at the time of LPS (3.88, 2.46-6.11), hemorrhagic infarct (1.94, 1.19-3.15), male gender (1.79, 1.10-2.92) and hyperglycemia (1.62, 1.05-2.51). CONCLUSIONS: In young ischaemic stroke patients, the magnitude of seizure risk and the major risk factors were similar to older ischaemic stroke patients but risk factors for ASSs and LPSs differed.

Cerebral hemodynamics in human acute ischemic stroke: a study with diffusion- and perfusion-weighted magnetic resonance imaging and SPECT.

Nineteen patients with acute ischemic stroke (<24 hours) underwent diffusion-weighted and perfusion-weighted (PWI) magnetic resonance imaging at the acute stage and 1 week later. Eleven patients also underwent technetium-99m ethyl cysteinate dimer single-photon emission computed tomography (SPECT) at the acute stage. Relative (ischemic vs. contralateral control) cerebral blood flow (relCBF), relative cerebral blood volume, and relative mean transit time were measured in the ischemic core, in the area of infarct growth, and in the eventually viable ischemic tissue on PWI maps. The relCBF was also measured from SPECT. There was a curvilinear relationship between the relCBF measured from PWI and SPECT (r = 0.854; P < 0.001). The tissue proceeding to infarction during the follow-up had significantly lower initial CBF and cerebral blood volume values on PWI maps (P < 0.001) than the eventually viable ischemic tissue had. The best value for discriminating the area of infarct growth from the eventually viable ischemic tissue was 48% for PWI relCBF and 87% for PWI relative cerebral blood volume. Combined diffusion and perfusion-weighted imaging enables one to detect hemodynamically different subregions inside the initial perfusion abnormality. Tissue survival may be different in these subregions and may be predicted.

The phorbol derivatives thymeleatoxin and 12-deoxyphorbol-13-O-phenylacetate-10-acetate cause translocation and down-regulation of multiple protein kinase C isozymes.

Phorbol esters such as phorbol 12-myristate,13-acetate (PMA) are potent activators of protein kinase C (PKC), and activate all PKC isozymes except zeta and lambda. Recently, 12-deoxyphorbol-13-O-phenylacetate-20-acetate (dPPA) and thymeleatoxin (Tx) were reported to selectively activate PKC beta 1 (dPPA) and PKC alpha, -beta, and -gamma (Tx), but not PKC delta or PKC epsilon in vitro. We examined the ability of these phorbol derivatives to translocate and down-regulate PKC isozymes in intact cells. Our findings demonstrate that dPPA and Tx cause translocation and down-regulation of multiple PKC isozymes, including delta and epsilon.

Protein kinase C-beta-like immunoreactivity and phosphorylation dependent immunoreactivity of neurofilaments in developing, adult and aged rat peripheral neurons.

The localizations of protein kinase C-beta-like immunoreactivity (PKC-beta-LI) and phosphorylated neurofilament-like immunoreactivity (PNF-LI) were studied in developing (E12-P28), adult (3-month-old) and aged (15- and 30-month-old) rat dorsal root (DRG) and superior cervical (SCG) ganglia. A close correlation was found between the localization of PKC-beta-LI and PNF-LI in all age groups. PKC-beta-LI was already present in the rat DRG at the time of the appearance of PNF-LI (E12) and after E15 both PKC-beta-and PNF-LI were found in the same subpopulation of neuronal perikarya. Aging had no effect on the distribution or intensity of either PKC-beta or PNF-LI. In the SCG neurons no association was observed between immunoreactivity and accumulation of lipopigments, but in the DRG neurons, lipopigment accumulation was seen in PKC-beta and PNF-immunoreactive large neurons. The results suggest that PKC-beta may regulate the time of appearance and phosphorylation state of some forms of phosphorylated neurofilaments during development. The results also show that no major change occurs in the amount or distribution of PNFs during normal aging.

Differential expression of immediate early genes in the superior cervical ganglion after nicotine treatment.

We examined the effects of single and multiple systemic injections of nicotine on the expression of five immediate early genes in the rat superior cervical ganglion by in situ hybridization histochemistry. A single nicotine injection resulted in a rapid and transient activation phase of nerve growth factor I-A, c-fos and jun-B at 20 min, and a later and less prominent activation of c-jun, which stayed high from 20 to 60 min. there was a parallel slow and long-lasting activation of jun-D, which remained high 4 h after nicotine treatment. Tyrosine hydroxylase mRNA, but not neuropeptide Y mRNA, was also induced by nicotine. Denervation of the ganglion did not prevent the induction of immediate early genes, but the nicotine antagonists hexamethonium and mecamylamine completely blocked the induction of immediate early genes, indicating that nicotine acted directly on receptors present on ganglion cells. When repeated nicotine injections were given, there was a refractory period of 1-2 h for c-fos, nerve growth factor I-A and jun-B induction. Repeated nicotine injections at 1-h intervals prevented about 80% of c-fos, nerve growth factor I-A and jun-B mRNA induction seen after a single injection. Because nicotine is known to induce immediate early genes in the adrenal glands as well, we examined whether similar kinetics of the gene induction could be seen in the adrenal medulla. However, no refractory period for repeated nicotine treatment or down regulation of the induction of the immediate early genes could be demonstrated in the adrenal medulla. The results show that sympathetic neurons respond to nicotine with altered expression of immediate early genes. Nicotine-induced expression of immediate early genes may be mediated and regulated by different factors in neuronal and endocrine noradrenergic cells.

Spreading depression induces expression of calcium-independent protein kinase C subspecies in ischaemia-sensitive cortical layers: regulation by N-methyl-D-aspartate receptors and glucocorticoids.

Spreading depression is a wave of sustained depolarization challenging the energy metabolism of the cells without causing irreversible damage. In the ischaemic brain, sreading depression-like depolarization contributes to the evolution of ischaemia to infarction. The depolarization is propagated by activation of N-methyl-D-aspartate receptors, but changes in signal transduction downstream of the receptors are not known. Because protein phosphorylation is a general mechanism whereby most cellular processes are regulated, and inhibition of N-methyl-D-aspartate receptors or protein kinase C is neuroprotective, the expression of protein kinase C subspecies in spreading depression was examined. Cortical treatment with KCl induced an upregulation of protein kinase Cdelta and zeta messenger RNA at 4 and 8 h, whereas protein kinase Calpha, beta, gamma and epsilon did not show significant changes. The gene induction was the strongest in layers 2 and 3, and was followed by an increased number of protein kinase Cdelta-immunoreactive neurons. Protein kinase Cdelta and zeta inductions were inhibited by pretreatment with an N-methyl-D-aspartate receptor antagonist, dizocilpine maleate, which also blocked spreading depression propagation, and with dexamethasone, which acted without blocking the propagation. Quinacrine, a phospholipase A2 inhibitor, reduced only protein kinase C5 induction. In addition, N(G)(-nitro-L-arginine methyl ester, a nitric oxide synthase inhibitor, did not influence protein kinase Cdelta or zeta induction, whereas 6-nitro-7-sulphamoylbenzo[f]quinoxaline-2,3-dione, an alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate/kainate receptor antagonist, and the cyclo-oxygenase inhibitors indomethacin and diclophenac tended to increase gene expression. The data show that cortical spreading depression induces Ca2(+)-independent protein kinase C subspecies delta and zeta, but not Ca(2+)-dependent subspecies, through activation of N-methyl-D-aspartate receptors and phospholipase A2. Even though the signal pathway is similar to the induction described previously in ischaemia for genes implicated in delayed neuronal death, the gene inductions observed here are not necessarily pathogenetic, but may represent a general reaction to metabolic stress.

Combined SPECT and diffusion-weighted MRI as a predictor of infarct growth in acute ischemic stroke.

UNLABELLED: In acute ischemic stroke, the infarcted core is surrounded by a zone of tissue that has decreased perfusion. Some of this tissue may be salvaged by prompt, effective treatment. Diffusion-weighted MRI is sensitive in detecting the infarcted tissue, whereas SPECT also detects the hypoperfused tissue around the infarcted core. We studied the potential of combined diffusion-weighted MRI and SPECT to predict infarct growth and clinical outcome in patients not receiving thrombolytic treatment. METHODS: Sixteen patients with acute stroke were examined consecutively with diffusion-weighted MRI and 99mTc-ethyl cysteinate dimer (99mTc-ECD) SPECT within 24 h of the onset of symptoms. Follow-up diffusion-weighted MRI was performed on the second day and after 1 wk. The volumes of infarcted and hypoperfused brain tissue were measured from diffusion-weighted MRI and SPECT, respectively. The volume difference between the hypoperfused and infarcted tissue on the first day was compared with the possible increase in infarct volume during the follow-up. Each patient's neurologic status was assessed with the National Institutes of Health Stroke Scale (NIHSS). RESULTS: The volume of infarcted tissue increased from 48 +/- 54 cm3 (mean +/- SD) on the first day to 88 +/- 93 cm3 on the second day (P = 0.001) and to 110 +/- 121 cm3 at 1 wk (P = 0.001). The volume of hypoperfused tissue on the first day was significantly greater than the infarct volume (102 +/- 135 cm3; P = 0.001). The volume difference between the hypoperfused and infarcted tissue on the first day correlated significantly with the infarct growth between the first day and 1 wk (r = 0.71; P < 0.01). Between the first day and 1 wk, the increase of the infarct volume correlated significantly with the change in the NIHSS (r = 0.54; P < 0.05). CONCLUSION: A large hypoperfusion zone around the infarct core in the acute phase of ischemic stroke predicts the infarct growth during the first week, and this correlates significantly with the change in the neurologic status of the patient. Combined diffusion-weighted MRI and SPECT performed within 24 h after the onset of symptoms can be useful in the evaluation of acute stroke to predict infarct growth.

Protein kinase C and adaptation to ethanol.

Adaptation to chronic ethanol exposure results in a decrease in sensitivity to the intoxicating effects of ethanol. Recent evidence indicates that changes in the expression and function of certain proteins involved in signal transduction are important for adaptation to ethanol. Using the neural cell line PC12, we found that chronic exposure to ethanol increases the expression and function of L-type voltage-gated calcium channels and enhances neural differentiation induced by nerve growth factor. Both of these responses to ethanol require protein kinase C (PKC). Chronic ethanol exposure activates PKC-mediated phosphorylation, in part, by increasing the expression of two PKC isozymes, delta and epsilon. The PKC family of enzymes may be important targets for the development of drugs that could modify adaptive and toxic consequences of chronic ethanol exposure.

Circadian rhythm in c-fos protein expression in the rat adrenal cortex.

The circadian variation in the expression of c-fos protein(s) in the adrenal cortex of adult rats was investigated immunocytochemically. The animals were maintained in standard laboratory facilities with a 12:12 h light:dark schedule (07.00-19.00 h) until sacrificed. The number of c-fos immunoreactive (c-fos IR) cells was statistically constant in the zona fasciculata and zona reticulata at 20.00 h and 24.00 h, and 04.00 h, but was decreased to 52% at 08.00 h and to 18% at 12.00 h, respectively, when compared to the 24.00 h value. At 16.00 h, the number of c-fos IR cells was increased again to 63% of the 24.00 h value. In the zona granulosa c-fos IR was observed only at 20.00 h and 24.00 h. Administration of dexamethasone, a potent inhibitor of ACTH release, significantly reduced the number of c-fos IR cells. As the circadian rhythm of c-fos expression in the adrenal cortex correlates to that reported for ACTH secretion from the pituitary, the results suggest that c-fos gene is involved in mediating physiological ACTH-induced responses in the adrenal cortical cells.

Histochemical detection of age- and injury-related changes in signal transduction in the superior cervical ganglion.

The superior cervical ganglion (SCG) is thought to be a good model for correlation studies of morphology, function and metabolism of neurons. The SCG has a relatively simple organization, it can be easily manipulated in situ, and it maintains synaptic transmission and a high metabolic rate during in vitro incubations. The histology and structure of SCG neurons have been characterized in detail, and physiologic stimuli, injury and aging have all been found to induce changes in the SCG morphology. During the last decade, research in the field of signal transduction has greatly expanded. Several signal transduction pathways have been identified that participate in the regulation of neurotransmitter synthesis, gene expression, neuronal excitability and growth factor responses of sympathetic neurons. We have been interested in using the SCG to study some of the second and third messengers involved in converting external stimuli received by sympathetic neurons into cellular short- and long-term events. Using immunohistochemistry, we have investigated protein kinase C-subtypes and the immediate early gene product Fos in the SCG, and characterized some of the changes induced by injury and aging in these messenger molecules. We will review the results and discuss the advantages and disadvantages of using histological methods in the study of signal transduction in sympathetic neurons.

Injury-induced long-term expression of immediate early genes in the rat superior cervical ganglion.

Using in situ hybridization histochemistry and immunocytochemistry the induction of immediate early genes (IEGs) was studied in the rat superior cervical ganglion (SCG) after deafferentation, axotomy and sialectomy (removal of the submandibular gland). Control SCGs showed very low levels of IEGs, whereby Fos proteins were found in 1% and Jun protein in 6% of neurons, but not in non-neuronal cells. Denervation and axotomy induced c-fos, NGFI-A, c-jun, jun B and jun D mRNA expression for up to 6 days in non-neuronal cells, whereas in sympathetic neurons the expression of only c-jun mRNA was induced after axotomy and sialectomy. The induction of Fos and Jun proteins by neuronal injury was demonstrated by immunocytochemistry. The results indicate that neuronal injury induces IEGs mainly in non-neuronal cells and that in neurons only c-jun is induced after axotomy. It is hypothesized that the induction of IEG other than c-jun in neurons after brain injury is an indirect event unrelated to the neuronal response to injury.

Increase in protein kinase-C-beta-like immunoreactivity (PKC-beta-LI) in the rat superior cervical ganglion after decentralization.

The effect of decentralization on protein kinase-C-beta-like immunoreactivity (PKC-beta-LI) in the rat superior cervical ganglion (SCG) was studied. The cervical sympathetic trunk was transected and at different time points (0 h-14 d) after transection the rats were decapitated and the SCGs examined immunohistochemically. In the control ganglia only faint PKC-beta-LI was seen in the principal neurons, whereas several strongly immunolabelled nerve fibers were observed. In the principal ganglionic neurons PKC-beta-LI was found to be increased 1-8 days after denervation, with the maximum accumulation occurring at 4 days. Fourteen days post-transection PKC-beta-LI in the neuronal perikarya was back to the initial level, but a decrease in the number of PKC-beta-immunoreactive nerve fibers was observed. The results suggest that the amount of PKC-beta in the principal neurons of the rat SCG may be partly regulated transsynaptically by preganglionic innervation.

Localization of protein kinase C-beta-like immunoreactivity in the rat dorsal root ganglion.

The localization of protein kinase C-beta-like immunoreactivity (PKC-beta-LI) was studied in the rat dorsal root ganglion (DRG) using an antibody specific for a peptide sequence common to the beta 1- and beta 2-subtypes. PKC-beta-LI was seen in 45% of neuronal cell bodies and in nerve fibers, which were mostly myelinated. The PKC-beta-LI-containing cell bodies had a diameter significantly larger than the unlabeled cell bodies. The results suggest that PKC-beta is a PKC subtype involved in cell surface signal transduction in the subpopulation of large DRG neurons.

Protein kinase C isozymes that mediate enhancement of neurite outgrowth by ethanol and phorbol esters in PC12 cells.

Using PC12 cells to study ethanol's effects on growth of neural processes, we found that ethanol enhances NGF- and basic FGF-induced neurite outgrowth. Chronic ethanol exposure selectively up-regulates delta and epsilon protein kinase C (PKC) and increases PKC-mediated phosphorylation in PC12 cells. Since PKC regulates differentiation, we investigated the role of PKC in enhancement of neurite outgrowth by ethanol. Like ethanol, 0.3-10 nM phorbol 12-myristate, 13-acetate (PMA) increased NGF-induced neurite outgrowth. However, higher concentrations did not, and immunoblot analysis demonstrated that 100 nM PMA markedly depleted cells of beta, delta and epsilon PKC. PMA (100 nM) also down-regulated beta, delta and epsilon PKC in ethanol-treated cells and completely prevented enhancement of neurite outgrowth by ethanol. In contrast, the cAMP analogue 8-bromoadenosine cAMP did not completely mimic the effects of ethanol on neurite outgrowth, and ethanol was able to enhance neurite formation in mutant PC12 cells deficient in protein kinase A (PKA). These findings implicate beta, delta or epsilon PKC, but not PKA, in the neurite-promoting effects of ethanol and PMA. Since chronic ethanol exposure up-regulates delta and epsilon, but not beta PKC, these findings suggest that delta or epsilon PKC regulate neurite outgrowth.

Decentralization induces long-term c-fos protein-like immunoreactivity in non-neuronal cells in the rat superior cervical ganglion.

The expression of c-fos protein(s) was studied immunohistochemically in the intact and decentralized superior cervical ganglia (SCG) of adult rats. C-fos protein(s)-like immunoreactivity (c-fos-LI) was detected in 1-2% of the principal ganglion cells but not in the satellite or Schwann cells of the intact SCG. After transection of the cervical sympathetic trunk, satellite and Schwann cells in the ganglion expressed c-fos-LI from one hour through 8 days following the operation. The maximal staining was seen 1 day postransection (d.p.t.), when the c-fos-LI was intense throughout the ganglion. No change was observed in the c-fos-LI of the principal neurons. At 8 d.p.t. the number of immunoreactive cells and their staining intensity had decreased, and at 14 d.p.t. they were back to the control level. These results suggest that a long-term expression of c-fos protein(s) is induced in the SCG satellite and Schwann cells after preganglionic denervation possibly related to their activation during nerve degeneration and regeneration.

Immunohistochemical evidence for the presence of protein kinase C beta-subtype in rat motoneurons and myoneural junctions.

The localization of protein kinase C beta-subtype-like immunoreactivity (PKC-beta-LI) was studied in the spinal cord and in different striated muscles of rat. In the spinal cord, large motoneurons in the ventral horn were PKC-beta-immunoreactive (IR). Strong immunoreactivity to PKC-beta was found in large nerve bundles between muscles, and in smaller nerves among muscle fibers. Myoneural junctions, which showed weaker immunoreactivity to PKC-beta, were also demonstrated in all muscles studied; the external ocular muscles, the diaphragma and the triceps surae muscle. Muscle cells were not labelled.

Early serial SPET in acute middle cerebral artery infarction.

The size and severity of perfusion defects in acute cerebral ischaemia on single photon emission tomographic (SPET) images may provide useful information regarding long-term (> 3 month) stroke outcome. A decreased predictive value has been reported with delayed SPET more than 24 h after stroke onset. We examined 20 patients with acute middle cerebral artery (MCA) infarctions using serial 99Tcm-ECD or 99Tcm-HMPAO SPET (SPET 1 one day and SPET 2 three days after stroke onset). Neurological (NIH, SSS) and functional (Barthel, Rankin) scores were calculated simultaneously and 3 months poststroke. The two SPET scans correlated equally well with the severity of functional and neurological deficits evaluated 3 months after stroke onset. In comparison to clinical assessment, the prognostic value of SPET was relatively better on the first day than the third day. Crossed cerebellar diaschisis correlated with early SPET deficits, but did not predict functional outcome. Our results suggest that SPET, either with 99Tcm-ECD or 99Tcm-HMPAO, can be used to predict stroke outcome in acute MCA infarction up to 72 h poststroke without significant interference from luxury perfusion.

Diffusion and perfusion MR imaging in acute ischemic stroke: a comparison to SPECT.

Diffusion (DWI) and perfusion (PWI) magnetic resonance imaging are relatively new methods of clinical imaging that probably can detect infarcted (DWI) and hypoperfused but still salvageable tissue (PWI) in acute human stroke. Forty-six acute stroke patients were imaged within 24 h of ictus, on the second day and after a week. SPECT was also performed on 23 patients in the acute phase (first or second day). On the first day, mean volume of hypoperfused tissue was significantly greater (P<0.001) than the infarcted tissue. The initial hypoperfusion volume correlated significantly with the final infarct size (P<0.001). The initial perfusion-diffusion mismatch correlated significantly with the infarct growth (P< or =0.001). The hypoperfusion volumes measured from PWI and SPECT correlated significantly (P<0.001). In conclusion, combined DWI and PWI is a powerful tool in evaluating the hemodynamics of acute ischemic stroke and can predict the infarct growth during 1 week.

Assessing the outcome of stroke: a comparison between MRI and clinical stroke scales.

OBJECTIVES: To assess the correlation of diffusion-weighted (DWI) and perfusion-weighted imaging (PWI) findings with the severity of acute neurologic deficit and their ability to predict short and long-term clinical outcomes of stroke. The ability of DWI and PWI to predict the outcome was compared with the ability of clinical stroke scales to predict the outcome. METHODS: Forty-eight patients with acute stroke underwent diffusion DWI and PWI on the first and eighth day after the ictus. Clinical and functional scales were carried out before each scan and 3 months after the stroke. RESULTS: The volumes of both the DWI and the PWI lesions correlated well with the acute neurologic deficit and the final outcome. The first day PWI (r = 0.64) and the National Institutes of Health Stroke Scale (NIHSS) scores (r = 0.70) correlated well with the final outcome. However, in logistic regression analysis, only the NIHSS score at the acute stage was the only independent predictor of the long-term clinical outcome. CONCLUSION: While the PWI and DWI lesion volumes correlated well with the outcome of the stroke, the imaging measurements did not improve the prognostic power over plain clinical stroke scale scores.