Retroviral Vector Stability: Inactivation Kinetics and Membrane Properties

2005 ◽  
pp. 303-308 ◽  
Author(s):  
P.E. Cruz ◽  
M. Carmo ◽  
A.S. Coroadinha ◽  
A. Bengala ◽  
D. Gonçalves ◽  
...  
Keyword(s):  
Retroviral Vector ◽  
Membrane Properties ◽  
Inactivation Kinetics

Na+, K+ and Ca2+ currents in identified leech neurones in culture

1989 ◽  
Vol 141 (1) ◽  
pp. 1-20
Author(s):  
R. R. Stewart ◽  
J. G. Nicholls ◽  
W. B. Adams
Keyword(s):  
Minor Component ◽  
Outward Current ◽  
Membrane Properties ◽  
Delayed Rectifier ◽  
Inactivation Kinetics ◽  
Channel Blockers ◽  
Sensory Neurones ◽  
K Current ◽  
The Individual ◽  
K Currents

1. Na+, K+ and Ca2+ currents have been measured by voltage-clamp in Retzius (R), anterior pagoda (AP) and sensory (pressure, touch and nociceptive) cells dissected from the central nervous system (CNS) of the leech. These cells maintain their distinctive membrane properties and action potential configurations in culture. Currents carried by the individual ions were analysed by the use of channel blockers and by their kinetics. Since the cells are isopotential they can be voltage-clamped effectively. 2. Depolarization, as expected, gave rise to an early inward Na+ current followed by a delayed outward K+ current. In Na+-free medium containing tetraethylammonium (TEA+), and in the presence of 4-aminopyridine (4-AP), inward Ca2+ currents were revealed that inactivated slowly and were blocked by Cd2+ and Mn2+. 3. Na+ and Ca2+ currents were similar in their characteristics in R. AP and sensory neurones. In contrast, K+ currents showed marked differences. Three principal K+ currents were identified. These differed in their time courses of activation and inactivation and in their responses to Ca2+ channel blockers. 4. K+ currents of the A-type (IA) activated and inactivated rapidly, were not affected by Ca2+ channel blockers and were eliminated by steady-state inactivation at holding potentials of −30 mV. A-type K+ currents were found in AP cells and as a minor component of the outward current in R cells. A Ca2+-activated K+ current (IC), that inactivated more slowly and was reduced by Ca2+ channel blockers, constituted the major outward current in R cells. The third K+ current resembled the delayed rectifier currents (IK1 and IK2) of squid axons with slow activation and inactivation kinetics. Such currents were found in R cells and in the sensory neurones (T, P and N). 5. The principal differences in membrane properties of identified leech neurones can be explained in terms of the numbers of Na+ channels and the distinctive kinetics of K+ channels in each type of cell.

Persistent Sodium Current, Membrane Properties and Bursting Behavior of Pre-Bötzinger Complex Inspiratory Neurons In Vitro

2002 ◽  
Vol 88 (5) ◽  
pp. 2242-2250 ◽  
Author(s):  
Christopher A. Del Negro ◽  
Naohiro Koshiya ◽  
Robert J. Butera ◽  
Jeffrey C. Smith
Keyword(s):  
Sodium Current ◽  
Neonatal Rat ◽  
Membrane Properties ◽  
Negative Slope ◽  
Inactivation Kinetics ◽  
Pacemaker Neurons ◽  
Inspiratory Neurons ◽  
Voltage Ramp ◽  
Bötzinger Complex

We measured persistent Na+current and membrane properties of bursting-pacemaker and nonbursting inspiratory neurons of the neonatal rat pre-Bötzinger complex (pre-BötC) in brain stem slice preparations with a rhythmically active respiratory network in vitro. In whole-cell recordings, slow voltage ramps (≤100 mV/s) inactivated the fast, spike-generating Na+ current and yielded N-shaped current-voltage relationships with nonmonotonic, negative-slope regions between −60 and −35 mV when the voltage-sensitive component was isolated. The underlying current was a TTX-sensitive persistent Na+ current ( I NaP) since the inward current was present at slow voltage ramp speeds (3.3–100 mV/s) and the current was blocked by 1 μM TTX. We measured the biophysical properties of I NaP after subtracting the voltage-insensitive “leak” current ( I Leak) in the presence of Cd2+ and in some cases tetraethylammonium (TEA). Peak I NaP ranged from −50 to −200 pA at a membrane potential of −30 mV. Decreasing the speed of the voltage ramp caused time-dependent I NaPinactivation, but this current was present at ramp speeds as low as 3.3 mV/s. I NaP activated at −60 mV and obtained half-maximal activation near −40 mV. The subthreshold voltage dependence and slow inactivation kinetics of I NaP, which closely resemble those of I NaP mathematically modeled as a burst-generation mechanism in pacemaker neurons of the pre-BötC, suggest that I NaP predominantly influences bursting dynamics of pre-BötC inspiratory pacemaker neurons in vitro. We also found that the ratio of persistent Na+conductance to leak conductance ( g NaP/ g Leak) can distinguish the phenotypic subpopulations of bursting pacemaker and nonbursting inspiratory neurons: pacemaker neurons showed g NaP/ g Leak> g NaP/ g Leakin nonpacemaker cells ( P < 0.0002). We conclude that I NaP is ubiquitously expressed by pre-BötC inspiratory neurons and that bursting pacemaker behavior within the heterogeneous population of inspiratory neurons is achieved with specific ratios of these two conductances, g NaP and g Leak.

scholarly journals Effect of Extracellular HCO3 − on Na+Channel Characteristics in Hippocampal CA1 Neurons

2000 ◽  
Vol 84 (5) ◽  
pp. 2477-2483 ◽  
Author(s):  
Xiang Q. Gu ◽  
Hang Yao ◽  
Gabriel G. Haddad
Keyword(s):  
Membrane Properties ◽  
Inactivation Kinetics ◽  
Na Channel ◽  
Action Potential Firing ◽  
Patch Clamp Technique ◽  
Ca1 Neurons ◽  
Hippocampal Ca1 Neurons ◽  
Neuronal Membranes ◽  
Recovery From Inactivation ◽  
Hippocampal Ca1

The effect of HCO3 −/CO2 on membrane properties of isolated hippocampal CA1 neurons was studied with the use of the whole cell configuration of the patch-clamp technique. Neurons were acutely dissociated from 21- to 30-day-old mice. In the current-clamp mode, HCO3 −/CO2significantly hyperpolarized CA1 neurons by more than 10 mV and decreased their input resistance. In addition, the overall excitability of these neurons was lower in the presence of HCO3 −/CO2 than in HEPES. Spontaneous and evoked action potential firing frequency was lower in the presence of HCO3 −/CO2 than in its absence. In the voltage-clamp mode, both activation and steady-state inactivation of a fast Na+ current were shifted in the hyperpolarized direction in such a way that the window currents were smaller in HCO3 −/CO2 than in HEPES. Recovery from inactivation and deactivation from the open state of the fast Na+ current was slower in HCO3 −/CO2 than in HEPES. We conclude that HCO3 −/CO2decreases the intrinsic excitability of CA1 neurons by altering not only the passive properties of the neuronal membranes but also by changing several characteristics of the fast Na+current, including activation and inactivation kinetics as well as the recovery from inactivation and deactivation.

Relationship between retroviral vector membrane and vector stability

2006 ◽  
Vol 87 (5) ◽  
pp. 1349-1356 ◽  
Author(s):  
M. Carmo ◽  
T. Q. Faria ◽  
H. Falk ◽  
A. S. Coroadinha ◽  
M. Teixeira ◽  
...  
Keyword(s):  
Chemical Properties ◽  
Mouse Skin ◽  
Retroviral Vectors ◽  
Retroviral Vector ◽  
Medium Composition ◽  
Transduction Efficiency ◽  
Inactivation Kinetics ◽  
Membrane Composition ◽  
Scanning Calorimetry ◽  
Physico Chemical

The present work studies the physico-chemical properties of retroviral vector membrane, in order to provide some explanation for the inactivation kinetics of these vectors and to devise new ways of improving transduction efficiency. For this purpose, vectors with an amphotropic envelope produced by TE Fly A7 cells at two culture temperatures (37 and 32 °C) were characterized by different techniques. Electron paramagnetic resonance (EPR) results showed that vectors produced at 32 °C are more rigid than those produced at 37 °C. Further characterization of vector membrane composition allowed us to conclude that the vector inactivation rate increases with elevated cholesterol to phospholipid ratio. Differential scanning calorimetry (DSC) showed that production temperature also affects the conformation of the membrane proteins. Transduction studies using HCT116 cells and tri-dimensional organ cultures of mouse skin showed that vectors produced at 37 °C have higher stability and thus higher transduction efficiency in gene therapy relevant cells as compared with vectors produced at 32 °C. Overall, vectors produced at 37 °C show an increased stability at temperatures below 4 °C. Since vector membrane physico-chemical properties are affected in response to changes in culture temperature, such changes, along with alterations in medium composition, can be used prospectively to improve the stability and the transduction efficiency of retroviral vectors for therapeutic purposes.

Physicochemical study of basement membrane properties iii. diffusion of proline, glucose and insulin in krebs solution

1988 ◽  
Vol 85 ◽  
pp. 391-397
Author(s):  
Bo Tao Fan ◽  
Françoise Simonnet ◽  
Jean Schaeverbeke ◽  
Gérard Lapluye
Keyword(s):  
Basement Membrane ◽  
Membrane Properties ◽  
Physicochemical Study ◽  
Krebs Solution
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