Sodium-dependent regenerative responses in dendrites of axotomized motoneurons in the cat.

223

127

Spinal sensory (dorsal root ganglion; DRG) neurons display slowly inactivating, tetrodotoxin-resistant (TTX-R), and rapidly inactivating, TTX-sensitive (TTX-S) Na currents. Attenuation of the TTX-R Na current and enhancement of TTX-S Na current have been demonstrated in cutaneous afferent DRG neurons in the adult rat after axotomy and may underlie abnormal bursting. We show here that steady-state levels of transcripts encoding the ␣-SNS subunit, which is associated with a slowly inactivating, TTX-R current when expressed in oocytes, are reduced significantly 5 days following axotomy of DRG neurons, and continue to be expressed at reduced levels, even after 210 days. Steady-state levels of ␣-III transcripts, which are present at low levels in control DRG neurons, show a pattern of transiently increased expression. In situ hybridization using ␣-SNS-and ␣-IIIspecific riboprobes showed a decreased signal for ␣-SNS, and an increased signal for ␣-III, in both large and small DRG neurons following axotomy. Reduced levels of ␣-SNS may explain the selective loss of slowly inactivating, TTX-R current. The abnormal electrophysiological properties of DRG neurons following axonal injury thus appear to ref lect a switch in Na channel gene expression.Multiple voltage-gated Na currents with different kinetics and voltage dependence have been observed in spinal sensory (dorsal root ganglion; DRG) neurons, which relay signals from peripheral receptors into the central nervous system. These include rapidly inactivating, TTX-sensitive (TTX-S) currents, and slowly inactivating, TTX-resistant (TTX-R) Na currents (1-4). Interplay of the currents influences the firing properties of DRG neurons (5-8) and may contribute to ectopic generation of action potentials in these cells following injury to their axons within peripheral nerves (9).Neural Na channels are heterotrimers composed of an ␣ subunit and two ␤ subunits, ␤1 and ␤2 (10 -12). Nucleotide sequence variation allows identification of ␣ subunits within single neurons by in situ hybridization (ISH) or in a cDNA pool using reverse transcriptase-PCR (RT-PCR) and restriction enzyme polymorphism (REP). ISH and RT-PCR͞ REP analyses have demonstrated that transcripts of multiple channel ␣ subunits, including ␣-I, ␣-II, NaG, Na6, and hNE (PN1), are expressed in single DRG neurons (13). The expression of these transcripts may explain the diversity of Na currents in these cells. Recently, a novel ␣ subunit , also termed PN3 (15)] was identified and found to be restricted to peripheral sensory neurons. ␣-SNS contains a serine at a site within domain I which has been shown, by site-directed mutagenesis, to confer resistance to TTX in the cardiac muscle Na channel (16). Expression of ␣-SNS cRNA produces a slowly inactivating, TTX-R current in Xenopus oocytes, suggesting that ␣-SNS is responsible for a TTX-R Na current in DRG neurons (14,15).Altered neuronal excitability has been demonstrated following axonal transection (17-19) and may underlie abnormal DRG bursting associated with pain...

Section: Discussioncontrasting

confidence: 37%

mentioning

confidence: 99%

Down-regulation of transcripts for Na channel α-SNS in spinal sensory neurons following axotomy

Dib‐Hajj

Black²,

Felts

et al. 1996

223

127

“…Since dendritic and axonal amputations have occurred in our dissociated neurons, a redistribution of Na+ channels onto dendrites, similar to that shown in axotomized vagal neurons (17), might be proposed to explain these results. It is well known that axotomy can affect Na+-dependent action tOther data show that there is a progressive increase in the density of Na+ currents in pyramidal neurons from embryonic day 16 to P19 (14).…”

Section: Discussionmentioning

confidence: 58%

Sodium channels in dendrites of rat cortical pyramidal neurons.

Huguenard

Hamill

Prince

1989

155

The voltage-dependent properties that have been directly demonstrated in Purkinje cell and hippocampal pyramidal cell dendrites play an important role in the integrative capacities of these neurons. By contrast, the properties of neocortical pyramidal cell dendritic membranes have been more difficult to assess. Active dendritic conductances near sites of synaptic input would have an important effect on the input-output characteristics of these neurons. In the experiments reported here, we obtained direct evidence for the existence of voltage-dependent Na+ channels on the dendrites of neocortical neurons by using cell-attached patch and whole cell recordings from acutely isolated rat neocortical pyramidal cells. The qualitative and quantitative properties of dendritic and somatic currents were indistinguishable. Insofar as Na+ currents are concerned, the soma and primary apical dendrite can be considered as one relatively uniform compartment. Similar dendritic Na+ currents on dendrites in mature neurons would play an important role in determining the integrative properties of these cortical units.Contrary to earlier concepts (1), recent data suggest that dendritic membranes of neurons in the mammalian cerebellum (2) and hippocampus (3-5) possess voltage-dependent conductances that lead to spike generation and play an important role in the integrative activities ofthese cells (2, 6). Evidence of dendritic spike generation in cortical pyramidal neurons is less complete. It has been proposed that lowamplitude, short-duration spikes seen in recordings from these cells in mature and immature neocortex are generated by voltage-dependent dendritic processes (7,8), although other explanations are possible (9). However, because of the fine caliber of neocortical pyramidal cell dendrites and the absence of a discrete dendritic layer, the anatomically confirmed, high-quality intradendritic recordings necessary to further evaluate membrane electroresponsiveness have been largely unavailable (10). We have used acutely isolated cortical neurons (11,12) and patch clamp techniques (13) to obtain direct evidence for the presence of voltage-dependent Na+ channels on the dendrites of rat neocortical pyramidal cells and have determined that, within the age range studied [postnatal days 2-6 (P2-P6)], the density and voltage dependence of Na+ channels on somatic and primary dendritic membrane are comparable. MATERIALS AND METHODSNeocortical pyramidal cells were isolated by using a combination of enzymatic and mechanical techniques (14) based on the methods of Wong and coworkers (11,12). Rat pups aged P2-P6 were anesthetized by cooling and then were decapitated. Brains were quickly removed, and 600-,m coronal slices containing sensorimotor cortex were cut on a vibratome (Lancer). Pieces of neocortex (1 mm2) were trimmed, incubated in a solution containing trypsin at 1 mg/ml, and then mechanically dispersed by trituration with fire-polished Pasteur pipettes.Pyramidal neurons in isolation (e.g., Figs. 1A, 2C, and 3A) were identif...

“…Excitability and Na current density are altered in neurons after axonal injury (22)(23)(24)(25). After axotomy, rat DRG neurons display dramatic changes in their TTX-R and TTX-S Na currents and in their Na channel mRNA profile; these changes include an attenuation of TTX-R and enhancement of TTX-S Na currents (21, 26), down-regulation of SNS͞PN3 transcripts and up-regulation of ␣III transcripts (20), and moderate elevation in the levels of ␣I and ␣II mRNAs (27).…”

Section: Introductionmentioning

confidence: 99%

NaN, a novel voltage-gated Na channel, is expressed preferentially in peripheral sensory neurons and down-regulated after axotomy

Dib‐Hajj

Tyrrell

Black

et al. 1998

469

306

Although physiological and pharmacological evidence suggests the presence of multiple tetrodotoxinresistant (TTX-R) Na channels in neurons of peripheral nervous system ganglia, only one, SNS͞PN3, has been identified in these cells to date. We have identified and sequenced a novel Na channel ␣-subunit (NaN), predicted to be TTX-R and voltage-gated, that is expressed preferentially in sensory neurons within dorsal root ganglia (DRG) and trigeminal ganglia. The predicted amino acid sequence of NaN can be aligned with the predicted structure of known Na channel ␣-subunits; all relevant landmark sequences, including positively charged S4 and pore-lining SS1-SS2 segments, and the inactivation tripeptide IFM, are present at predicted positions. However, NaN exhibits only 42-53% similarity to other mammalian Na channels, including SNS͞PN3, indicating that it is a novel channel, and suggesting that it may represent a third subfamily of Na channels. NaN transcript levels are reduced significantly 7 days post axotomy in DRG neurons, consistent with previous findings of a reduction in TTX-R Na currents. The preferential expression of NaN in DRG and trigeminal ganglia and the reduction of NaN mRNA levels in DRG after axonal injury suggest that NaN, together with SNS͞PN3, may produce TTX-R currents in peripheral sensory neurons and may inf luence the generation of electrical activity in these cells.Voltage-gated Na channels in rat brain are composed of three subunits: the pore-forming ␣-subunit (260 kDa), which is sufficient to generate Na current flow across the membrane, and two auxiliary subunits, ␤1 (36 kDa) and ␤2 (33 kDa), which can modulate the properties of the ␣-subunit (1, 2). Nine distinct ␣-subunits have been identified in the rat (ref. 3 and references therein, refs. 4-6), and homologues have been cloned from various mammalian species, including humans (3). Specific ␣-subunits are expressed in a tissue-and developmentally specific manner (7). Aberrant expression patterns or mutations of voltage-gated sodium channel ␣-subunits underlie a number of human and animal disorders (1, 8-11).Multiple voltage-gated Na currents, some tetrodotoxinsensitive (TTX-S) and others TTX-resistant (TTX-R) (12-15), have been observed in dorsal root ganglia (DRG) neurons, which express multiple Na channel ␣-subunit mRNAs (16). The complex Na current profile in DRG neurons influences their excitability (13,(17)(18)(19) and may contribute to ectopic or spontaneous firing in these cells after injury to their axons (8,20,21).Excitability and Na current density are altered in neurons after axonal injury (22)(23)(24)(25). After axotomy, rat DRG neurons display dramatic changes in their TTX-R and TTX-S Na currents and in their Na channel mRNA profile; these changes include an attenuation of TTX-R and enhancement of TTX-S Na currents (21, 26), down-regulation of SNS͞PN3 transcripts and up-regulation of ␣III transcripts (20), and moderate elevation in the levels of ␣I and ␣II mRNAs (27). Inflammatory modulators also up-regulate TTX-R c...