Vesicular storage of glycine in glutamatergic terminals in mouse hippocampus.

Glycine acts as a neuromodulator to regions rich in glutamatergic synapses, such as the forebrain. However, recent evidences for synaptic release of glycine in hippocampal cultured neurons and synaptosomes argue for the existence of functional glycinergic synapses in the hippocampus. It is well established that GABA and glycine act in concert at inhibitory synapses, while the existence of synapses which utilize both glutamate and glycine is less common. The purpose of the present study was to investigate the distribution of glycine and its role in hippocampal neurotransmission. Using immunohistochemistry, we demonstrate that vesicular glycine is preferentially stored in glutamatergic, rather than GABAergic presynaptic terminals. Using the sniffer patch technique, we found that glycine could be released upon presynaptic activity. Furthermore, using whole-cell patch-clamp recordings, we show for the first time the presence of a postsynaptic strychnine-sensitive chloride current in response to presynaptic stimulation. The small amplitude of this current is likely due to the paucity of postsynaptic glycine receptors rather than a low level of glycine release. Taken together, our results suggest that glycine is stored in glutamatergic presynaptic terminals. It is likely that the major role of glycine that is released from presynaptic terminals is to modulate N-methyl-d-aspartate receptor function but may also play a role in decreasing neuronal excitability by opposing glutamatergic neurotransmission in pathological states such as epilepsy or ischemia.

P73 regulates cisplatin-induced apoptosis in ovarian cancer cells via a calcium/calpain-dependent mechanism.

P73 is important in drug-induced apoptosis in some cancer cells, yet its role in the regulation of chemosensitivity in ovarian cancer (OVCA) is poorly understood. Furthermore, if and how the deregulation of p73-mediated apoptosis confers resistance to cisplatin (CDDP) treatment is unclear. Here we demonstrate that TAp73α over-expression enhanced CDDP-induced PARP cleavage and apoptosis in both chemosensitive (OV2008 and A2780s) and their resistant counterparts (C13* and A2780cp) and another chemoresistant OVCA cells (Hey); in contrast, the effect of ΔNp73α over-expression was variable. P73α downregulation attenuated CDDP-induced PUMA and NOXA upregulation and apoptosis in OV2008 cells. CDDP decreased p73α steady-state protein levels in OV2008, but not in C13*, although the mRNA expression was identical. CDDP-induced p73α downregulation was mediated by a calpain-dependent pathway. CDDP induced calpain activation and enhanced its cytoplasmic interaction and co-localization with p73α in OV2008, but not C13* cells. CDDP increased the intracellular calcium concentration ([Ca(2+)](i)) in OV2008 but not C13* whereas cyclopiazonic acid (CPA), a Ca(2+)-ATPase inhibitor, caused this response and calpain activation, p73α processing and apoptosis in both cell types. CDDP-induced [Ca(2+)](i) increase in OV2008 cells was not effected by the elimination of extracellular Ca(2+), but this was attenuated by the depletion of internal Ca(2+) store, indicating that mobilization of intracellular Ca(2+]) stores was potentially involved. These findings demonstrate that p73α and its regulation by the Ca(2+)-mediated calpain pathway are involved in CDDP-induced apoptosis in OVCA cells and that dysregulation of Ca(2+)/calpain/p73 signaling may in part be the pathophysiology of CDDP resistance. Understanding the cellular and molecular mechanisms of chemoresistance will direct the development of effective strategies for the treatment of chemoresistant OVCA.

Do-not-resuscitate decision: the attitudes of medical and non-medical students.

OBJECTIVES: To study the attitudes of both medical and non-medical students towards the do-not-resuscitate (DNR) decision in a university in Hong Kong, and the factors affecting their attitudes. METHODS: A questionnaire-based survey conducted in the campus of a university in Hong Kong. Preferences and priorities of participants on cardiopulmonary resuscitation in various situations and case scenarios, experience of death and dying, prior knowledge of DNR and basic demographic data were evaluated. RESULTS: A total of 766 students participated in the study. There were statistically significant differences in their DNR decisions in various situations between medical and non-medical students, clinical and preclinical students, and between students who had previously experienced death and dying and those who had not. A prior knowledge of DNR significantly affected DNR decision, although 66.4% of non-medical students and 18.7% of medical students had never heard of DNR. 74% of participants from both medical and non-medical fields considered the patient's own wish as the most important factor that the healthcare team should consider when making DNR decisions. Family wishes might not be decisive on the choice of DNR. CONCLUSIONS: Students in medical and non-medical fields held different views on DNR. A majority of participants considered the patient's own wish as most important in DNR decisions. Family wishes were considered less important than the patient's own wishes.

Total intravenous anaesthesia with propofol and remifentanil for elective neurosurgical procedures: an audit of early postoperative complications.

BACKGROUND AND OBJECTIVES: This was a prospective audit to assess the incidence and characteristics of early postoperative complications in the recovery room in extubated patients after elective neurosurgical procedures using propofol and remifentanil-based total intravenous anaesthesia. METHODS: Vital signs (temperature, conscious level, respiratory rate, oxygen saturation, pulse and blood pressure) and postoperative complications (shivering, nausea, vomiting and cardiorespiratory) were analysed in 145 adult patients over a 1-yr period. RESULTS: The overall shivering, postoperative nausea and vomiting and postoperative hypertension (systolic blood pressure more than 25% of the preoperative value) incidences were 30.3%, 16.6% and 35.2%, respectively. Fifty-one percent of the patients had at least one of the above complications. The complication rates were found to be widely different among various types of neurosurgery. The surgical procedures were divided into five groups: supratentorial craniotomy, posterior fossa craniotomy, intracranial vascular procedures, transphenoidal hypophysectomy and extracranial procedures. Median extubation time was similar in all groups and patients were fully conscious with no hypoxia in the recovery room. The intracranial vascular group had the highest shivering and postoperative nausea and vomiting rates (58.8% and 29.4%, respectively). In the supratentorial craniotomy group, 46% of the patients had hypertension. The overall complication rate (presence of any complications) was highest in the supratentfial craniotomy (55.4%), posterior fossa craniotomy (75%) and intracranial vascular (76.5%) groups. Shivering and overall complication rate was significantly related to the anaesthetic time (P

Large venous air embolism in the sitting position despite monitoring with transoesophageal echocardiography.

A 49-year-old male with neurofibromatosis type II was scheduled for posterior fossa craniotomy and excision of a right acoustic neuroma and placement of an auditory brainstem implant in the sitting position. Intra-operatively, the patient was monitored with transoesophageal echocardiography which detected two major episodes of venous air embolism. Despite immediate treatment the patient's gas exchange progressively worsened during surgery and a chest X-ray showed extensive bilateral pulmonary infiltrates. The patient developed acute respiratory distress syndrome and required inotropic support in the intensive care unit. Although transoesophageal echocardiography allowed rapid detection of venous air embolism, there was no evidence of therapeutic benefit.

Detecting synaptic connections in the medial nucleus of the trapezoid body using calcium imaging.

The study of synaptic transmission in brain slices generally entails the patch-clamping of postsynaptic neurones and stimulation of identified presynaptic axons using a remote electrical stimulating electrode. Although patch recording from postsynaptic neurones is routine, many presynaptic axons take tortuous turns and are severed in the slicing procedure, blocking propagation of the action potential to the synaptic terminal and preventing synaptic stimulation. Here we demonstrate a method of using calcium imaging to select postsynaptic cells with functional synaptic inputs prior to patch-clamp recording. We have used this method for exploring transmission in the auditory brainstem at the medial nucleus of the trapezoid body neurones, which are innervated by axons from the contralateral cochlear nucleus. Brainstem slices were briefly loaded with the calcium indicator fura-2 AM and stimulated with an electrode placed on the midline. Electrical stimulation caused a rise in intracellular calcium concentration in those postsynaptic neurones with active synaptic connections. Since <10% of the medial nucleus of the trapezoid body neurones retain viable synaptic inputs following the slicing procedure, preselecting those cells with active synapses dramatically increased our recording success. This detection method will greatly ease the study of synaptic responses in brain areas where suprathreshold synaptic inputs occur but connectivity is sparse.