Ultrastructural detection of neuronally transported choleragenoid by postembedding immunocytochemistry in freeze-substituted Lowicryl HM20™ embedded tissue

Choleragenoid (cholera toxin B-fragment; CTB) is an anterograde, retrograde and transganglionic neuronal tracer. We describe a method for detecting CTB-labeled neuronal cell bodies, neurites and boutons at the ultrastructural level, using postembedding immunogold techniques on freeze-substituted Lowicryl HM20™ embedded nervous tissue. Primary afferents and motoneurons were labeled by injection of CTB in the dorsal ramus of the C2 spinal nerve of the rat. Following fixation with paraformaldehyde (4%) and glutaraldehyde (0.25%), tissue sections from the spinal cord C2 segment were freeze-substituted and embedded in Lowicryl HM20™ and subsequently processed with postembedding immunocytochemistry for CTB and glutamate. Immunogold particles indicating CTB immunoreactivity were found over primary afferents and motoneurons. In primary afferents in the central cervical nucleus (CCN) and motor nuclei, immunogold labeling was seen in boutons over vesicle-containing axoplasm and to a lesser extent over axoplasm devoid of vesicles, but not over mitochondria or axolemma. In motoneurons, immunogold particles were seen over the Golgi apparatus in the soma and over lysosomes in both soma and dendrites. Quantification of glutamate-like immunoreactivity in 20 CTB-labeled and 20 CTB-negative boutons in the neuropil was found similar, indicating that CTB does not interfere with the immunocytochemical detection of neuronal epitopes such as the transmitter substance glutamate.     

Chronic treatment with SCH-23390, a selective dopamine D1 receptor blocker decreases preprotachykinin-A mRNA levels in nucleus tractus solitarii of the rabbit: role in respiratory control.

Acute intravenous administration of the selective D1 receptor blocker SCH-23390 resulted in an enhanced respiratory motor output as evidenced by the phrenic nerve activity, whereas local perfusion into the region of nucleus tractus solitarii had no effect. The increase in phrenic nerve activity was accompanied by a concomitant increase in the release of substance P in the region of nucleus tractus solitarii as measured by in vivo microdialysis technique. Chronic administration of SCH-23390 via subcutaneously implanted Alzet mini osmotic pumps, significantly decreased the level of preprotachykinin-A mRNA in the region of respiratory relay neurons in nucleus tractus solitarii but was without effect in the ventral medullary surface structure, wherein the central chemoreceptors are thought to be located. A smaller, but significant decrease was also seen in the striatum. The results suggest that chronic treatment with SCH-23390 leads to a disinhibition of an inhibitory dopaminergic input to the neurons in nucleus tractus solitarii from a suprapontine level, which may account for a subsequent inhibition of tachykinin-containing neurons in the nucleus tractus solitarii, the relay station for respiratory reflexes.     

Myocardial sestamibi single-photon emission tomography: Variations in reference values with gender, age and rest versus stress?

Reference data files support the evaluation of myocardial perfusion single-photon emission tomography (SPET). The aim of this study was to create a large reference data base for technetium-99m sestamibi SPET, age and gender matched to the general patient population. One hundred and twenty-eight healthy volunteers (76 males and 52 females) with a likelihood of coronary artery disease of less than 5% underwent rest and maximal exercise^99mTc-sestamibi SPET with a 2-day protocol and 180° elliptical rotation. The normalized activity values of^99mTc-sestamibi in the inferior wall differed significantly between men and women. Age variations were found for men in the anterior wall. Normalized activity values in all four walls were strikingly similar during rest and stress. Our results suggest that the use of reference files in^99mTc-sestamibi SPET requires a gender- and, for males, possibly an age-matched reference population. Different reference files at rest and during stress might not be necessary.     

Sensory neuropeptides contribute to oedema formation in experimental burns

The role of primary sensory neuropeptides in experimental burns was investigated. It was shown that after scalding, substance P-like immunoreactivity is released into the lymphatic fluid of dogs. Substance P, injected intravenously, was shown to cause dose-dependent oedema formation in rat skin, as measured with the Evans blue-technique. Furthermore, capsaicin pretreatment, which depletes the neuropeptide content of primary sensory neurons, was shown to inhibit late oedema formation in rats.     

Effects of cooling and heating of the tooth on pulpal blood flow in man

Abstract Laser Doppler flowmetry (LDF) was used to study the changes in pulpal blood flow (PBF) evoked by application of cold or heat to the palatinal surfaces of teeth 11 or 21 in nine young subjects. Switching from a thermode temperature of 33° to 5° G on average induced a slow decrease of PBF to about 80% of control, and also warming to 39°C evoked a small reduction in most subjects. Inter individual differences were large, however, and both cooling and warming sometimes triggered a rise in PBF. In contrast, skin blood flow, as recorded with LDF in the forearm, invariably rose during warming and fell during local cooling. The results suggested a more complex interaction between local and nervously mediated effects of moderate changes in temperature in the tooth pulp than skin, and that the previous held view of cold and heat decreasing and increasing PBF, respectively, is wrong.     

Effects of posture and pneumoperitoneum during anaesthesia on the indices of left ventricular filling

Background. Laparoscopic surgery requires the use of pneumoperitoneum (PP). When combined with positional changes, pneumoperitoneum may cause marked circulatory alterations.
Methods. Eight anaesthetized cardiovascularly healthy patients, scheduled for laparoscopic cholecystectomy, were studied before and during pneumoperitoneum in three different postures (supine, Trendelenburg and reversed Trendelenburg), employing transesophageal echocardiography and pulmonary artery pressure monitoring.
Results. PP significantly increased end-diastolic area (EDA) and pulmonary capillary wedge pressure (PCWP) irrespective of posture. PCWP was significantly influenced by postural changes, whereas EDA was not. Further, changes in EDA and PCWP covaried during the investigation, but showed no linear correlation. Systolic function, measured as end-systolic area (ESA) and fractional area shortening (FAS), was not altered. Diastolic function, as assessed by the velocity rate of the trans-mitral flow during the early filling phase (E) and the atrial contraction (A), showed no change of the E/A ratio, whereas after the induction of PP there was a significant reduction of the E component.
Conclusions. In cardiovascularly healthy patients, the left ventricular volume is increased during pneumoperitoneum. Further, changes in invasive pressure determinations (PCWP) do not correlate linearly with changes in volume indices of left ventricular filling (EDA).     

Effect of dipyridamole (Persantin) on blood flow and patency of aortocoronary vein bypass grafts

The effect of dipyridamole was investigated in 360 patients undergoing coronary bypass surgery. They were randomly allocated to receive dipyridamole (100 mg orally q.i.d. for 2 days preoperatively, 5 mg/kg body weight/24 h i.v. peroperatively and 100 mg orally q.i.d. for 1 year postoperatively) or placebo. Withdrawn from the study were 48 patients on dipyridamole and 57 on placebo. Cardiovascular and/or cerebrovascular events or need for anticoagulant treatment were the reasons for withdrawal in 22 (13%) of the dipyridamole, and 34 (18%) of the placebo group. Logistic regression analysis of risk factors influencing graft patency showed significant relation to peroperatively measured coronary blood flow. A positive trend of treatment was observed (p = 0.08). Vein graft blood flow measured during bypass surgery (245 patients) was significantly greater in the dipyridamole group (p less than 0.01). The occlusion rate was lower in vessels with peroperative blood flow greater than 30 ml/min (vein-marginal p less than 0.01, vein-dexter p less than 0.05, vein-diagonal 0.05 less than p less than 0.1). Dipyridamole increases coronary blood flow and graft patency following coronary bypass surgery.