Prolonged Blood Storage Does Not Effect Survival in an Animal Model of Hemorrhagic Shock.

SUMMARY: BACKGROUND: Red blood cell (RBC) transfusion in hemorrhagic shock is life saving. However, several clinical trials have shown that blood transfusion in the critically ill patient might be associated with adverse outcomes. Furthermore, an association between prolonged blood storage and adverse effects of RBC transfusion has been postulated. The aim of this study is to examine the effect of blood storage time on resuscitation outcome, in an animal model of hemorrhagic shock. METHODS: 20 Wistar rats were phlebotomized in order to induce reversible hemorrhagic shock. Half of them were resuscitated with blood stored for a short period of time (4 days), and the other ones were resuscitated with blood stored for a prolonged time (14 days). Blood samples for hemoglobin, pH, lactate, bicarbonate and creatinine were drawn prior to the induction of shock and 24 h after resuscitation. Five days after resuscitation the animals were sacrificed, and liver, lung and kidney histology was examined. RESULTS: At 24 h after bleeding, the hemoglobin levels decreased by 3.2 and 1.7 g/dl, the pH decreased by 0.008 and 0.001, while the lactate levels increased by 1.6 and 2.7 mg/dl in the fresh and old blood resuscitation groups, respectively, with no significant difference between the groups. A trend toward more severe renal damage occurred in the old compared to the fresh blood resuscitation group (p = 0.089). CONCLUSION: The results of the present study show that in this animal model of hemorrhagic shock the duration of storage of RBCs used for transfusion did not affect the outcome of resuscitation.

Spatiotemporal aspects of pulsed electrical stimuli on the responses of rabbit retinal ganglion cells.

Implanted intraocular microelectrode arrays are being used to provide sight to individuals who are blind due to photoreceptor degeneration. It is envisioned that this retinal prosthesis will create the illusion of motion by stimulating focal areas of the retina in a sequential fashion through neighboring electrodes, much like the rapid succession of still images in movies and computer animation gives rise to apparent motion. Using a high-density microelectrode array, we examined the extracellularly recorded responses of rabbit retinal ganglion cells to a bar-shaped electrode array that was stepped at 50 microm increments at different rates across the retina and compared these responses to the responses generated to a similarly shaped light stimulus that was stepped across the retina. When the retina was stimulated at 1 step/s, retinal ganglion cells gave robust bursts of action potentials to both the electrode array and the light stimulus. The responses to the 'moving' electrode array decreased progressively with increasing stepping frequency. At 16 steps/s (highest frequency tested), the number of spikes per sweep and the number of bursts per sweep were reduced 75% and 67% respectively. In contrast, when the retina was stimulated at 16 steps/s with the 'moving' light stimulus, the number of spikes per sweep and the number of bursts per sweep were reduced only 43% and 25% respectively. These findings suggest that simple translation of object motion to sequential stimulation through neighboring electrodes may not be the best way to convey the perception of object motion in a patient with a retinal prosthesis.

Thresholds for activation of rabbit retinal ganglion cells with relatively large, extracellular microelectrodes.

PURPOSE: To investigate the responses of retinal ganglion cells (RGCs) to electrical stimulation, using electrodes comparable in size to those used in human studies investigating the feasibility of an electronic retinal prosthesis. METHODS: Rabbit retinas were stimulated in vitro with current pulses applied to the inner surface with 125- and 500-mum diameter electrodes while the responses of RGCs were recorded extracellularly. RESULTS: Both short-latency (SL; 3-5 ms) and long-latency (LL; >/=9 ms) responses were observed after electrical stimulation within the receptive field of an RGC. With short, 0.1-ms current pulses, the threshold current for the SL cell response was significantly lower than that for the LL cell response. With long (10- to 20-ms) pulses, the threshold currents for the SL and LL cell responses were very similar. The threshold current for the SL cell response increased more steeply than did the LL cell response when the electrode was displaced from the point of lowest electrical threshold, either above or along the surface of the retina. Stimulation of an RGC axon outside of the cell's receptive field produced only an SL response. For 0.1-ms duration pulses, the threshold current for the axonal response was significantly higher than the threshold current for the SL cell response. At pulse durations > 1 ms, the thresholds were very similar. CONCLUSIONS: RGC responses to electrical stimulation depend on the current pulse duration and location of the stimulating electrode. For an epiretinal prosthesis, short-duration current pulses may be preferable since they result in a more localized activation of the retina.

Responses of rabbit retinal ganglion cells to electrical stimulation with an epiretinal electrode.

Rational selection of electrical stimulus parameters for an electronic retinal prosthesis requires knowledge of the electrophysiological responses of retinal neurons to electrical stimuli. In this study, we examined the effects of cathodal and anodal current pulses on the extracellularly recorded responses of OFF and ON rabbit retinal ganglion cells (RGCs) in an in vitro preparation. Current pulses (1 msec duration), delivered by a 125 microm electrode placed on the inner retinal surface within the receptive field of a RGC, produced both short-latency (< or =5 msec) and long-latency (8-60 msec) responses. The long-latency responses, but not the short-latency responses, were abolished upon application of the glutamate receptor antagonists CNQX and NBQX, thus indicating that the long-latency responses of RGCs are due to activation of presynaptic neurons in the retina. The latency of the long-latency response depended upon the polarity of the stimulus. For OFF RGCs, the average latency was 11 msec for a cathodal stimulus and 24 msec for an anodal stimulus. For ON RGCs, the average latency was 25 msec for a cathodal stimulus and 16 msec for an anodal stimulus. The threshold current also depended upon the polarity of the stimulus, at least for OFF RGCs. The average threshold current for evoking a long-latency response in OFF RGCs was 10 microA for a cathodal stimulus and 21 microA for an anodal stimulus. In ON RGCs, the average threshold current was 13 microA for a cathodal stimulus and 15 microA for an anodal stimulus.

In vitro activation of retinal cells: estimating location of stimulated cell by using a mathematical model.

Activation of neurons at different depths within the retina and at various eccentricities from the stimulating electrode will presumably influence the visual percepts created by a retinal prosthesis. With an electrical prosthesis, neurons will be activated in relation to the stimulating charge that impacts their cell membranes. The common model used to predict charge density is Coulomb's law, also known as the square law. We propose a modified model that can be used to predict neuronal depth that takes into account: (1) finite dimensions related to the position and size of the stimulating and return electrodes and (2) two-dimensional displacements of neurons with respect to the electrodes, two factors that are not considered in the square law model. We tested our model by using in vitro physiological threshold data that we had obtained previously for eight OFF-center brisk-transient rabbit retinal ganglion cells. For our most spatially dense threshold data (25 microm increments up to 100 microm from the cell body), our model estimated the depth of one RGC to be 76 +/- 76 microm versus 87 +/- 62 microm (median: SD) for the square law model, respectively. This difference was not statistically significant. For the seven other RGCs for which we had obtained threshold data up to 800 microm from the cell body, the estimate of the RGC depth (using data obtained along the X axis) was 96 +/- 74 versus 20 +/- 20 microm for the square law and our modified model, respectively. Although this difference was not statistically significant (Student t-test: p = 0.12), our model provided median values much closer to the estimated depth of these RGCs (>>25 microm). This more realistic estimate of cell depth predicted by our model is not unexpected in this latter data set because of the more spatially distributed threshold data points that were evaluated. Our model has theoretical advantages over the traditional square law model under certain conditions, especially when considering neurons that are horizontally displaced from the stimulating electrode. Our model would have to be tested with a larger threshold data pool to permit more conclusive statements about the relative value of our model versus the traditional square law model under special circumstances.

Human somatostatin receptor specificity of backbone-cyclic analogues containing novel sulfur building units.

Somatostatin-14 (somatostatin) and its clinically available analogues octreotide, lanreotide, and vapreotide are potent inhibitors of growth hormone, insulin, and glucagon release. Recently, a novel backbone cyclic somatostatin analogue c(GABA-Phe-Trp-(D)Trp-Lys-Thr-Phe-GlyC3-NH(2)) (analogue 1, PTR 3173) that possesses in vivo endocrine selectivity was described. This long-acting octapeptide exhibits high affinity to human recombinant somatostatin receptors (hsst) hsst2, hsst4, and hsst5. Its novel binding profile resulted in potent in vivo inhibition of growth hormone but not of insulin release. We report the synthesis, bioactivity, and structure-activity relationship studies of compounds related to 1. In these analogues, the lactam bridge of 1 was replaced by a backbone disulfide bridge. We present a novel approach for conformational constraint of peptides by utilizing sulfur-containing building units for on-resin backbone cyclization. These disulfide backbone cyclic analogues of 1 showed significant metabolic stability as tested in various enzyme mixtures. Receptor binding assays revealed different receptor selectivity profiles for these analogues in comparison to their prototype. It was found that analogues of 1, bearing a disulfide bridge, had increased selectivity to hsst2 and hsst5; however, they exhibited weaker affinity to hsst4 as compared to 1. These studies imply that ring chemistry, ring size, and ring position of the peptide template may affect the receptor binding selectivity.

Thresholds for activation of rabbit retinal ganglion cells with an ultrafine, extracellular microelectrode.

PURPOSE: To determine electrical thresholds required for extracellular activation of retinal ganglion cells as part of a project to develop an epiretinal prosthesis. METHODS: Retinal ganglion cells were recorded extracellularly in retinas isolated from adult New Zealand White rabbits. Electrical current pulses of 100- micro s duration were delivered to the inner surface of the retina from a 5- micro m long electrode. In about half of the cells, the point of lowest threshold was found by searching with anodal current pulses; in the other cells, cathodal current pulses were used. RESULTS: Threshold measurements were obtained near the cell bodies of 20 ganglion cells and near the axons of 19 ganglion cells. Both cathodal and anodal stimuli evoked a neural response in the ganglion cells that consisted of a single action potential of near-constant latency that persisted when retinal synaptic transmission was blocked with cadmium chloride. For cell bodies, but not axons, thresholds for both cathodal and anodal stimulation were dependent on the search method used to find the point of lowest threshold. With search and stimulation of matching polarity, cathodal stimuli evoked a ganglion cell response at lower currents (approximately one seventh to one tenth axonal threshold) than did anodal stimuli for both cell bodies and axons. With cathodal search and stimulation, cell body median thresholds were somewhat lower (approximately one half) than the axonal median thresholds. With anodal search and stimulation, cell body median thresholds were approximately the same as axonal median thresholds. CONCLUSIONS: The results suggest that cathodal stimulation should produce lower thresholds, more localized stimulation, and somewhat better selectivity for cell bodies over axons than would anodal stimulation.

Somatostatin analogue treatment attenuates histological findings of inflammation and increases mRNA expression of interleukin-1 beta in the articular tissues of rats with ongoing adjuvant-induced arthritis.

OBJECTIVE: Somatostatin is a neuropeptide with modulatory effects on the immune system and the function of synovial cells; it has antiangiogenic and antiproliferative properties. This study aimed to evaluate the clinical, histological, and articular tissue cytokine mRNA response to somostatin treatment in rat adjuvant-induced arthritis (AIA). METHODS: Adjuvant-induced arthritis was induced in a total of 68 Lewis rats by immunization with complete Freund's adjuvant. Twenty-four rats were treated with a long-acting somostatin analogue 14 days after disease induction. Twenty-four untreated rats served as controls. The severity of arthritis was scored weekly for 42 days. In a second experiment, 20 rats (ten treated, ten controls) were killed 21 days after treatment for assessment of joint histopathology and articular tissue cytokine mRNA expression. RESULTS: Somatostatin analogue treatment significantly reduced histological scores of early inflammatory changes and increased articular tissue mRNA expression of interleukin-1 beta (IL-1beta). A trend toward improvement in physical scores of joint inflammation was seen in the treated group. Late destructive changes were not significantly different. CONCLUSION: Treatment with a somostatin analogue attenuated early inflammatory changes in AIA joints and increased mRNA expression of IL-1beta in the articular tissues of rats with ongoing arthritis. Improvement in the physical findings of joint inflammation was mild.

Differential recovery of the electroretinogram, visually evoked cortical potential, and electrically evoked cortical potential following vitrectomy: implications for acute testing of an implanted retinal prosthesis.

To determine the extent to which electrophysiologic tests of the afferent visual pathway are affected by vitrectomy, the procedure was performed in 15 eyes of 11 adult Dutch-belted rabbits. An electroretinogram (ERG), visually evoked cortical potential (VECP), and electrically evoked cortical potential (EECP) were obtained preoperatively and sequentially after surgery. For electrical stimulations, biphasic impulses were delivered to the retina. Post-vitrectomy declines of 49, 25, and 41% from the median baseline amplitudes and increases of 13, 18, and 17% from the median baseline latency values were found for ERG, VECP, and EECP, respectively. At 90 min, 13 to 30% of eyes still had an amplitude more than 10% below baseline on at least one of the three tests, whereas 10 to 47% of eyes had an abnormal latency more than 10% above baseline on at least one of the three tests. Amplitudes were more likely than latencies to return to near baseline, but for eyes that remained subnormal, the decline was greater for amplitudes than latencies. Significant alterations in retinal function, manifested by declines in amplitudes and increases in latencies of the ERG, VECP, and EECP, persist in a large proportion of eyes up to 90 min post-vitrectomy.