mV In Neurons Research Articles

Dynamics of cell membrane lesions and adaptive conductance under the electrical stress.

Exceeding physiological limits of the cell membrane potential compromises structural integrity, enabling the passage of normally impermeant solutes and disrupting cell function. Electropermeabilization has been studied extensively at the cellular scale, but not at the individual membrane lesion level. We employed fast total internal reflection fluorescence (TIRF) imaging of Ca2+ entry transients to discern individual lesions in a hyperpolarized cell membrane and characterize their focality, thresholds, electrical conductance, and the lifecycle. A diffuse and momentary membrane permeabilization without a distinct pore formation was observed already at a -100 mV threshold. Polarizing down to -200 mV created focal pores with a low 50- to 300-pS conductance, which disappeared instantly once the hyperpolarization was removed. Charging to -240 mV created high-conductance (> 1 nS) pores which persisted for seconds even at zero membrane potential. With incremental hyperpolarization steps, persistent pores often emerged at locations different from those where the short-lived, low-conductance pores or diffuse permeabilization were previously observed. Attempts to polarize membrane beyond the threshold for the formation of persistent pores increased their conductance adaptively, preventing further potential build-up and "clamping" it at a certain limit (-270 ± 6 mV in HEK cells, -284 ± 5 mV in CHO cells, and -243 ± 9 mV in neurons). The data suggest a previously unknown role of electroporative lesions as a protective mechanism against a potentially fatal membrane overcharging and cell disintegration.

Deactivation of L-type Ca Current by Inhibition Controls LTP at Excitatory Synapses in the Cerebellar Nuclei

Long-term potentiation (LTP) of mossy fiber EPSCs in the cerebellar nuclei is controlled by synaptic inhibition from Purkinje neurons. EPSCs are potentiated by a sequence of excitation, inhibition, and disinhibition, raising the question of how these stimuli interact to induce plasticity. Here, we find that synaptic excitation, inhibition, and disinhibition couple to different calcium-dependent signaling pathways. In LTP induction protocols, constitutively active calcineurin can replace synaptic excitation, and constitutively active alpha-CaMKII can replace calcium influx associated with resumption of spiking upon disinhibition. Additionally, nimodipine can replace hyperpolarization, indicating that inhibition of firing decreases Ca influx through L-type Ca channels, providing a necessary signal for LTP. Together, these data suggest that potentiation develops after a calcineurin priming signal combines with an alpha-CaMKII triggering signal if and only if L-type Ca current is reduced. Thus, hyperpolarization induced by synaptic inhibition actively controls excitatory synaptic plasticity in the cerebellar nuclei.

Specific Kinetic Alterations of Human CaV2.1 Calcium Channels Produced by Mutation S218L Causing Familial Hemiplegic Migraine and Delayed Cerebral Edema and Coma after Minor Head Trauma

Mutation S218L in the Ca(V)2.1 alpha(1) subunit of P/Q-type Ca(2+) channels produces a severe clinical phenotype in which typical attacks of familial hemiplegic migraine (FHM) triggered by minor head trauma are followed, after a lucid interval, by deep (even fatal) coma and long lasting severe cerebral edema. We investigated the functional consequences of this mutation on human Ca(V)2.1 channels expressed in human embryonic kidney 293 cells and in neurons from Ca(V)2.1 alpha(1)(-/-) mice by combining single channel and whole cell patch clamp recordings. Mutation S218L produced a shift to lower voltages of the single channel activation curve and a consequent increase of both single channel and whole cell Ba(2+) influx in both neurons and human embryonic kidney 293 cells. Compared with the other FHM-1 mutants, the S218L shows one of the largest gains of function, especially for small depolarizations, which are insufficient to open the wild-type channel. S218L channels open at voltages close to the resting potential of many neurons. Moreover, the S218L mutation has unique effects on the kinetics of inactivation of the channel because it introduces a large component of current that inactivates very slowly, and it increases the rate of recovery from inactivation. During long depolarizations at voltages that are attained during cortical spreading depression, the extent of inactivation of the S218L channel is considerably smaller than that of the wild-type channel. We discuss how the unique combination of a particularly slow inactivation during cortical spreading depression and a particularly low threshold of channel activation might lead to delayed severe cerebral edema and coma after minor head trauma.

Differential alterations of the inactivation properties of high voltage activated calcium currents in area CA1 and CA3 of the rat following photothrombotic lesion

There are many indications that focal brain ischemia may alter the properties of remote brain tissue. We investigated whether changes of neuronal properties can be observed in the unlesioned ipsilateral hippocampus following cortical photothrombosis in the somatosensory cortex of rats. The whole-cell patch-clamp technique was used to investigate calcium current properties of hippocampal neurons (CA1 and CA3) 7 days after infarct induction. A significant alteration in the half-maximal potential of inactivation ( V h,i) could be demonstrated, when comparing lesioned with sham operated animals, while other current parameters remained unchanged. The alterations of the V h,i in the CA1 and CA3 regions were of opposite directions: V h,i in CA1 neurons was shifted negatively by 5.6 mV, and positively by 5.0 mV in neurons from the CA3 region. It has been speculated that these differential alterations may be due to different subunit compositions of calcium channels in these two brain areas. The data indicate that small cortical lesions can lead to widespread alterations of the neuronal network's excitability in the hippocampal formation.

Afferent Regulation of Inhibitory Synaptic Transmission in the Developing Auditory Midbrain

To determine whether afferent innervation regulates the strength of inhibitory connections in the gerbil auditory midbrain, both cochleas were surgically removed in postnatal day 7 animals, before sound-driven activity is first observed. Inhibitory synaptic currents were measured in a brain slice preparation 1-7 d after the ablations. Whole-cell and gramicidin-perforated patch recordings were obtained from inferior colliculus neurons, and IPSCs were evoked by stimulation of the commissure of the inferior colliculus (CIC) or the ipsilateral lateral lemniscus (LL) in the presence of kynurenic acid. Deafferentation led to a 24 mV depolarizing shift in the IPSC equilibrium potential within 1 d of deafferentation. As a consequence, there was a large reduction of IPSC amplitude at a holding potential of -20 mV in neurons from bilaterally ablated animals. Furthermore, both afferent pathways displayed a 50% reduction of the inhibitory synaptic conductance after deafferentation, indicating that driving force was not solely responsible for the decline in IPSC amplitude. When paired pulses were delivered to the LL or CIC pathway in control neurons, the evoked IPSCs exhibited facilitation. However, paired pulse facilitation was nearly eliminated after deafferentation. Thus, normal innervation affects inhibitory synaptic strength by regulating postsynaptic chloride homeostasis and presynaptic transmitter release properties.

Two components of muscarine-sensitive membrane current in rat sympathetic neurones.

Membrane currents induced by muscarine (Imus) were recorded in voltage-clamped neurones in isolated rat superior cervical ganglia. Two components of Imus were regularly recorded: an inward current resulting from inhibition of the outward K+ current, IM; and an outward current attributable to the reduction of a steady inward current. The presence of these two components caused a 'cross-over' in the current-voltage curves at -50 +/- 3 mV in neurones impaled with KCl-filled micro-electrodes or at -63 +/- 4 mV in neurones impaled with K-acetate-filled electrodes. Both components of Imus were prevented by atropine. Both persisted in Krebs solution containing tetrodotoxin (1 microM), Cd2+ (200 microM) or 0 Ca2+. When IM was inhibited by external Ba2+ or internal Cs+ only the outward component of Imus could be detected. This component reversed at +3 +/- 2 mV in cells impaled with CsCl-filled electrodes or at -20 +/- 3 mV in cells impaled with Cs-acetate-filled electrodes. The reversal potentials agreed with those for the currents induced by gamma-aminobutyric acid (+4 +/- 2 mV and -16 +/- 3 mV with CsCl and Cs acetate electrodes respectively). Replacement of external NaCl with Na acetate (so reducing external Cl- concentration ( [Cl-]o) from 155 to 22 mM) shifted the reversal potential for Imus by +25 and +14.5 mV in two cells impaled with CsCl-filled electrodes. A tenfold reduction of external [Na+] (by glucosamine replacement) did not significantly alter the reversal potential for Imus in KCl or CsCl-impaled cells. Under conditions where IM is already inhibited, the residual outward component of Imus can lead to hyperpolarization and inhibition of neuronal activity in unclamped cells. We conclude that both inward and outward components of Imus result from direct activation of muscarinic receptors on the ganglion cells. The inward component results from IM inhibition. We suggest that the outward component results from inhibition of another, voltage-independent current IX which largely comprises a Cl- current. The inward component induces membrane depolarization and an increased excitability; the outward component can lead to hyperpolarization and reduced excitability.

On the nature of histamine-mediated slow hyperpolarizing synaptic potentials in identified molluscan neurones.

1. Standard intracellular stimulating and recording techniques were used to test the correspondence between monosynaptic post-synaptic potentials (p.s.p.s) evoked by histamine-containing C-2 neurones and responses to focally applied histamine recorded from two classes of identified post-synaptic neurones in the cerebral ganglion of Aplysia californica.2. Two types of p.s.p.s were examined: (1) a monophasic slow hyperpolarizing potential (I(s)p.s.p.) lasting 1-2 sec; and (2) a biphasic p.s.p. consisting of a fast depolarizing component <0.5 sec in duration (E(f)p.s.p.) plus a slow hyperpolarizing potential (I(s)p.s.p.) designated the E(f)I(s)p.s.p.3. Ionophoretic or pressure applied histamine mimicked both p.s.p.s and produced conductance increases in the post-synaptic neurones similar to those associated with the evoked p.s.p.s.4. The reversal potentials (E(rev)) for the I(s)p.s.p. and E(f)I(s)p.s.p., estimated by extrapolation, were -85+/-5.3, -35+/-5.5, and -83+/-8.1 mV (mean+/-S.D.), respectively. The I(s)p.s.p.s were produced by an increase in potassium conductance because their E(rev)s were shifted about 16 mV by doubling or halving the concentration of extracellular potassium and they could be eliminated by cooling or by intracellular injection of TEA ions.5. The average E(rev) values for the slow hyperpolarizing histamine responses were similar to those for the I(s)p.s.p.s; about -83 and -86 mV in neurones receiving the monophasic I(s)p.s.p.s and biphasic E(f)I(s)p.s.p., respectively.6. Cimetidine, an antihistamine drug that selectively blocks histamine receptors associated with potassium conductances in Aplysia, reversibly abolished the I(s)p.s.p.s and slow hyperpolarizing responses to focally applied histamine. In similar concentrations, cimetidine had no discernible effects on the E(f)p.s.p. and depolarizing response to histamine or on several different types of p.s.p.s mediated by the C-2 neurones.7. It is proposed that the I(s)p.s.p.s are mediated by histamine released from the C-2 neurones.