Biomedical subjects

W Ma

Publications and source records attributed to W Ma.

At least 199 records · Page 11Linked to original sources

PubMed1998

Increased activation of nuclear factor kappa B in rat lumbar dorsal root ganglion neurons following partial sciatic nerve injuries.

Nuclear factor kappa B (NFkappaB) is a transcription factor which can be activated by some neurotrophic factors and cytokines, and then translocated into the nucleus. We examined NFkappaB immunoreactivity (IR) in L4 and L5 dorsal root ganglion (DRG) cells of normal rats, and 2 weeks after complete sciatic nerve transection (CSNT), partial sciatic nerve ligation (PSNL) and chronic constriction injury (CCI). In the normal DRG, 45% of the neurons were NFkappaB-IR (IR in cytoplasm only or in both cytoplasm and nucleus). Only 18% were activated NFkappaB-IR cells (IR in both cytoplasm and nucleus). Two weeks after CSNT, PSNL and CCI, there was no significant difference in the percentages of NFkappaB-IR neurons between the ipsilateral and contralateral DRG. However, the percentages of the activated NFkappaB-IR neurons in the ipsilateral DRG of PSNL (30%) and CCI (33%) rats, but not in CSNT (24%) rats, were significantly increased, compared with the contralateral DRG. Ultrastructurally, NFkappaB-IR was localized to the endoplasmic reticulum and Golgi apparatus. In activated cells, IR was also observed in the nuclei. Two weeks after CCI, NFkappaB-IR was stronger in the axons and Schwann cells in the proximal stump of the injured sciatic nerves than in uninjured contralateral nerves. In some Schwann cells surrounding unmyelinated fibers, the nuclei were also NFkappaB-IR, suggesting that these cells were activated by CCI. NFkappaB activation increased in DRG neurons and Schwann cells 2 weeks following partial sciatic injuries, possibly in response to cytokines and neurotrophins produced by endoneurial cells in the partially injured nerve during Wallerian degeneration.

Animals↗

PubMed1998

Phenanthroline-Cu complex-mediated chemiluminescence of DNA and its potential use in antioxidation evaluation.

The chemiluminescence (CL) concomitant with phen-Cu2+/ascorbate/H2O2-induced DNA damage has been studied. The emission intensity increases linearly with increasing DNA concentration. The emission spectrum has a maximal wavelength at about 410 nm. The luminescence is inhibited by histone in a histone concentration-dependent manner. The CL is characteristic of guanine. Of all common kinds of bases and nucleotides, only guanine or guanine nucleotides can give rise to luminescence. The possibility of using the luminescence as a means of studying antioxidation related to DNA damage is discussed. Several kinds of well-defined antioxidants have been used as testing reagents and show that this method can not only evaluate the antioxidative effect but also distinguish different types of antioxidants.

Antioxidants↗

PubMed1998

Cholesterol and bile acid metabolism are impaired in mice lacking the nuclear oxysterol receptor LXR alpha.

We demonstrate that mice lacking the oxysterol receptor, LXR alpha, lose their ability to respond normally to dietary cholesterol and are unable to tolerate any amount of cholesterol in excess of that which they synthesize de novo. When fed diets containing cholesterol, LXR alpha (-/-) mice fail to induce transcription of the gene encoding cholesterol 7alpha-hydroxylase (Cyp7a), the rate-limiting enzyme in bile acid synthesis. This defect is associated with a rapid accumulation of large amounts of cholesterol in the liver that eventually leads to impaired hepatic function. The regulation of several other crucial lipid metabolizing genes is also altered in LXR alpha (-/-) mice. These results demonstrate the existence of a physiologically significant feed-forward regulatory pathway for sterol metabolism and establish the role of LXR alpha as the major sensor of dietary cholesterol.

Alkyl and Aryl Transferases↗

PubMed1998

Extracellular ATP directly gates a cation-selective channel in rabbit airway ciliated epithelial cells.

1. A membrane conductance activated by extracellular ATP was identified and characterized in freshly dissociated rabbit airway ciliated cells using the whole-cell and outside-out patch configurations of the patch-clamp technique. 2. In solutions designed to maximize currents through voltage-gated calcium channels, there were no indications of voltage-gated Ba2+ currents. 3. Extracellular ATP (but not UTP or ADP) activated a membrane conductance which remained activated for several minutes in the presence of ATP. The conductance was permeable to monovalent and divalent cations with approximate relative permeabilities (P) for PBa : PCs : PTEA of 4 : 1 : 0.1. Permeability to Cl- was negligible. 4. Including GDP-beta-S in the intracellular solution did not inhibit the effects of ATP, nor did GTP-gamma-S irreversibly activate the conductance. 5. In outside-out membrane patches, with GDP-beta-S in the pipette solution, ATP activated ion channels which had a chord conductance of approximately 6 pS in symmetrical 150 mM CsCl solutions at -120 mV. 6. Suramin (100 microM) inhibited the whole-cell currents activated by ATP (200 microM) by 93 +/- 3 %. Similar effects of suramin were observed on ATP-activated channels in outside-out membrane patches. 7. Extracellular ATP had a priming action on the response to subsequent exposure to ATP. At -40 mV, the time to half-maximal current activation (t1/2) was 46 +/- 9 s during the first exposure to 200 microM ATP and decreased to 5 +/- 3 s during a second exposure to the same concentration of ATP. The priming action of ATP was not inhibited by including GDP-beta-S in the intracellular solution. 8. The initial rate of activation increased with the concentration of ATP, and was voltage sensitive. During the first exposure to 200 microM ATP, t1/2 at +40 mV was 4-fold longer than t1/2 at -40 mV. 9. Half-maximal activation of the conductance shifted from 210 +/- 30 to 14 +/- 4 microM added ATP when CaCl2 in the extracellular solution was reduced from 1.58 to 0. 01 mM. The Hill coefficient for ATP was 1.2 in both solutions.10. These observations suggest that a form of ATP uncomplexed with divalent cations directly gates an ion channel (P2X receptor) in rabbit airway ciliated cells, which serves as a pathway for Ca2+ influx. This purinoceptor may contribute to sustained ciliary activation during prolonged exposures to ATP.

Adenosine Diphosphate↗

PubMed1998

Neuronal and glial epitopes and transmitter-synthesizing enzymes appear in parallel with membrane excitability during neuroblastoma x glioma hybrid differentiation.

The membrane excitability and the presence of neural proteins, including neuronal and glial markers and neurotransmitter-synthesizing enzymes, were examined in parallel while the NG108-15 cell line was maintained in a serum-free medium. Whole-cell recordings in voltage-clamp or current-clamp configurations were used to evaluate the membrane excitability, and immunostaining was done with a panel of well-characterized antibodies against NSE, NF150, S-100 beta, GFAP, ChAT and TH. Culture for 4 to 10 days led to a striking rise in neurite outgrowth, electrical excitability and expression of neural proteins in type I neuron-like cells, which were of both neuronal and glial character, and expressed both cholinergic and adrenergic traits. After about 2 weeks, type II cells which lack neurite processes began to emerge. The type II cells proliferated, as revealed by BrdU uptake, and gradually overgrew differentiated cell types. They exhibited little or no membrane excitability and absence of immunoreactivity for the neuronal and glial specific proteins tested. These measurements indicate that the presence of these neural proteins at crucial stages of membrane excitability development is an important characteristics of NG108-15 cell differentiation, providing insights into the neural development and the reversible nature of neoplasia in the nervous system.

Animals↗

PubMed1998

GABA, GAD, and GABA(A) receptor alpha4, beta1, and gamma1 subunits are expressed in the late embryonic and early postnatal neocortical germinal matrix and coincide with gliogenesis.

Increasing evidence indicates that the classical, fast-acting neurotransmitter gamma-amino butyric acid (GABA) may initially act as morphogen in cell proliferation and differentiation via specific receptors. In view of the potential roles for GABA in central nervous system development, we examined the expression of GABA, GABA(A) receptor beta1 and gamma1 subunits by immunocytochemistry and the expression of transcripts for two GABA-synthesizing enzymes, glutamate decarboxylase (GAD65, GAD67 mRNAs), and for alpha4, beta1, and gamma1 subunits of GABA(A) receptor by in situ hybridization in the developing neocortex. Tissue sections were taken from embryonic days (E) 17 and E20 embryos and newborn rats (P0). The embryos' mothers and newborn rats had been injected with 5-bromo-2'-deoxyuridine (BrdU) and had survived for 2 hours. At E17, BrdU-positive cells were largely restricted in the synthetic zone at the ventricular margin when cortical neurogenesis was still active. GAD mRNAs and GABA immunoreactivity were detected in the subventricular zone, while alpha4, beta1, and gamma1 subunits were abundant in the ventricular zone. At E20 and P0, when neurogenesis had largely ceased and gliogenesis had commenced, BrdU-positive cells were found throughout the ventricular zone with GABA, GAD mRNAs, and alpha4, beta1, and gamma1 subunits. GABA, GAD mRNAs and alpha4, beta1, and gamma1 subunit signals intensified in the ventricular zone from E17 to P0 as gliogenesis proceeded. Thus, specific components of a putative GABAergic circuit are expressed in cells of the ventricular zone during the late embryonic/early postnatal period coincident with gliogenesis, suggesting a role for GABA in glial cell proliferation.

Animals↗

PubMed1998

Prenatal expression of inwardly rectifying potassium channel mRNA (Kir4.1) in rat brain.

The levels and cellular localization of the mRNA encoding the inwardly rectifying potassium ion channel Kir4.1 were investigated in the embryonic rat brain by Northern blots and in situ hybridization. This transcript was absent at embryonic day 13 (E13), whereas it was clearly present in E14-15 preparations, principally in the neuroepithelium of the cerebral cortex, thalamus, and hypothalamus. At later embryonic stages (E17-20), Kir4.1 mRNA levels increased and expanded to the mantle zone, such as the cortical plate, hippocampus, thalamus, and hypothalamus. The early appearance of Kir4.1 mRNA in various brain regions suggests an involvement of the channel in cell proliferation, migration and differentiation in the rat CNS.

Animals↗

PubMed1998

Basic FGF-responsive telencephalic precursor cells express functional GABA(A) receptor/Cl-channels in vitro.

We have previously described the expression of specific gamma-aminobutyric acid (GABA)A receptor subunits and their transcripts in the cortical neuroepithelium (Ma and Barker, 1995, 1998). However, it is not clear whether neural precursor cells exposed to basic fibroblast growth factor (bFGF) in vitro reproduce the biological properties of neuroepithelial cells in vivo within the embryonic ventricular zone. In the present study, neural precursor cells were isolated from the telencephalic neuroepithelium of embryonic day 13-13.5 rats and exposed to bFGF in serum-free medium. Basic FGF-responsive cells were capable of dividing and differentiating into neurons and astrocytes. The rapidly dividing cells formed multicellular spheres and then a rosette-like formation in which a majority of cells expressed GABA(A) receptor alpha4, beta1, or gamma1 subunit proteins. We found in perforated patch-clamp recordings that GABA depolarized bromodeoxyundine (BrdU)+ precursor cells, and under voltage-clamp induced a bicuculline-sensitive current that reversed at the Cl- equilibrium potential. GABA also increased cytoplasmic Ca2+ in a significant number of BrdU+ cells that was blocked by bicuculline. The bicuculline sensitivity of these pharmacological effects implicates GABA(A) receptor/Cl- channels, since bicuculline is a competitive GABA(A) antagonist at these channels in well-differentiated cells. It is possible that the three GABA(A) receptor subunits (alpha4, beta1, and gamma1) found in precursor cells form the Cl- channels detected electrophysiologically. The functional GABA(A) receptor/Cl- channels and associated regulation of their cytoplasmic Ca2+ levels via bicuculline-sensitive mechanisms may play significant roles in the regulation of neural cell proliferation in this model neuroepithelium.

Animals↗

PubMed1998

The effect of photochemical stress upon the lenses of normal and glutathione peroxidase-1 knockout mice.

This communication investigates the effect of oxidative stress upon the lenses of young normal and glutathione peroxidase-1 (GSHPx-1) Knockout mice. Both normal and knockout lenses have similar biochemical and morpholigical characteristics and the elimination of GSHPx-1 only decreases slightly the ability of the lens to degrade H2O2. Examination of the effect of a 4 hr photochemical stress on morphological characteristics indicates that there is comparable damage in the normal and knockout lenses in the epithelial and bow regions while the posterior region remains normal. However, at 24 hrs post-insult, the normal lenses appear to recover somewhat in the bow region while the knockout bow and posterior regions have extensive damage. In contrast to the morphological data, the biochemical parameters (14C)choline transport and (3H)thymidine incorporation are affected to a somewhat greater extent in the knockout lenses than in normal lenses. While both of these parameters are further affected in the 24 hr post-insult period, there is no further change in the relative effects upon normal and knockout lenses. Non-protein thiol is affected in a similar manner in both lens types. The effect upon biochemical parameters of tertiary butyl hydroperoxide (TBHP) insult was similar to H2O2 and photochemical stress. The overall conclusion is that young GSHPx-1 knockout lenses handle oxidative stress somewhat less effectively than comparable normal lenses but non-stressed knockout lenses appear normal. These results differ from observations reported by Reddy et al. (1997) under somewhat different conditions.

Animals↗

PubMed1998

The effect of catalase amplification on immortal lens epithelial cell lines.

Utilizing a human beta-actin promoter, a catalase cDNA expression vector was constructed. This construct was used to transfect two immortal cell lines, mouse alpha TN4-1 and rabbit N/N 1003A. The catalase activity was increased about 3.4 fold in the alpha TN4-1 cells and 38 fold in the N/N 1003A cells. Some changes in other enzyme activities were also observed as a result of the transfections. Surprisingly, the ability to degrade H2O2 in the extracellular environment of the cells did not markedly change as a result of the catalase amplification. However, the ability to resist H2O2 stress was dramatically altered. Non-protein thiol (NP-SH) levels, choline uptake and glyceraldehyde phosphate dehydrogenase (GPD) activity were all markedly decreased in the non-transfected cells when they were subjected to 300 microM H2O2. However, in both transfected cell lines, these parameters remained in the normal range during H2O2 stress. The results obtained upon observing aspects of DNA metabolism were more complicated. While on H2O2 stress, non-transfected cell lines showed a marked decrease in thymidine incorporation, only the transfected alpha TN4-1 line remained in the normal range. Thymidine incorporation in transfected rabbit N/N 1003A cells was decreased compared to normal cells. In contrast, studies on single strand DNA breaks indicated that transfected rabbit cells had little damage compared to the significant DNA damage observed in the normal cells. The normal N/N 1003A cells were also much more susceptible to H2O2 induced damage than normal alpha TN4-1 cells, suggesting that the high GSH peroxidase activity observed in the rabbit cells may be detrimental since the low glutathione reductase activity in such cells results in an accelerated depletion of glutathione. The overall results suggest that augmenting lens catalase may prevent cataract development caused by H2O2 stress.

Animals↗

PubMed1998

Increase of calcitonin gene-related peptide immunoreactivity in the axonal fibers of the gracile nuclei of adult and aged rats after complete and partial sciatic nerve injuries.

Neuropeptide changes in primary sensory neurons are thought to be involved in the pathological mechanisms of neuropathic pain caused by peripheral nerve injuries. In this study, using immunocytochemistry, we observed that calcitonin gene-related peptide (CGRP) immunoreactive (IR) fibers were increased, qualitatively and quantitatively, in the injured side gracile nuclei of adult (2 months old) and aged (16 months old) rats 2 weeks following complete transection (CSNT) or chronic constriction injury (CCI) of sciatic nerves. This increase was more pronounced after CCI than after CSNT. In aged rats, the CGRP-IR fibers which appeared were dystrophic. In contrast to the increases which we saw in the gracile nucleus, after both types of injury there was a decrease in CGRP-IR in all laminae of the dorsal horn. The percentage of CGRP-IR DRG neurons was decreased after CSNT, but unchanged after CCI. We interpret our results in terms of local sprouting in the gracile nucleus and suggest that the increased response following CCI is due to the involvement of fibers from DRG neurons spared by the partial nerve injury. Increased CGRP release from spared afferents in the gracile nucleus might be important in neuropathic pain.

Aging↗

PubMed1998

Partial and complete sciatic nerve injuries induce similar increases of neuropeptide Y and vasoactive intestinal peptide immunoreactivities in primary sensory neurons and their central projections.

Partial nerve injury is more likely to cause neuropathic pain than complete nerve injury. We have compared the changes in neuropeptide expression in primary sensory neurons which follow complete and partial injuries to determine if these might be involved. Since more neurons are damaged by complete injury, we expected that complete sciatic nerve injury would simply cause greater increases in neuropeptide Y and vasoactive intestinal peptide than partial injury. We examined neuropeptide Y and vasoactive intestinal peptide immunoreactivities in L4 and L5 dorsal root ganglia, the dorsal horn of L4-L5 spinal cord, and the gracile nuclei of rats killed 14 days after unilateral complete sciatic nerve transection, partial sciatic nerve transection and chronic constriction injury of the sciatic nerves. In all three groups of rats, neuropeptide Y- and vasoactive intestinal peptide-immunoreactive neurons were increased in the ipsilateral L4 and L5 dorsal root ganglion when compared with the contralateral side. Most neuropeptide Y-immunoreactive neurons were of medium and large size, but a few were small. Neuropeptide Y-immunoreactive axonal fibers were increased from laminae I to IV, and vasoactive intestinal peptide-immunoreactive axonal fibers were increased in laminae I and II, of the ipsilateral dorsal horn of L4-L5 spinal cord. The increases of neuropeptide Y and vasoactive intestinal peptide immunoreactivities in the dorsal horn were similar among the three groups. However, only after constriction injury were some vasoactive intestinal peptide-immunoreactive neurons seen in the deeper laminae of the ipsilateral dorsal horn. Robust neuropeptide Y-immunoreactive axonal fibers and some neuropeptide Y-immunoreactive cells were seen in the ipsilateral gracile nuclei of all three groups of animals, but neuropeptide Y-immunoreactive cells were more prominent after constriction injury. Contrary to our expectations, partial and complete sciatic nerve injuries induced similar increases in neuropeptide Y and vasoactive intestinal peptide in lumbar dorsal root ganglion neurons and their central projections in the dorsal horn and the gracile nuclei two weeks after injury. Some neurons whose axons were spared by partial injury may also increase neuropeptide Y or vasoactive intestinal peptide expression. Altered neuropeptide release from these functional sensory neurons may play a role in neuropathic pain.

Animals↗

PubMed1998

Molecular modeling study of tubulosine and other related ipecac alkaloids.

A molecular modeling study of two alkaloids, tubulosine and psychotrine, isolated from the sap of Pogonopus speciosus, and other related ipecac alkaloids, showed that these flexible alkaloids favor a nonplanar structure. The biologically active compounds had conformations with a similar angle between aromatic ring A, the nitrogen in ring B, and ring D. This angle was related to the biological activity reported for these compounds. Our results support the hypothesis of two different types of receptor interactions, one for the nonplanar compounds and another for the planar compounds.

Alkaloids↗

PubMed1998

Dominant GABA(A) receptor/Cl- channel kinetics correlate with the relative expressions of alpha2, alpha3, alpha5 and beta3 subunits in embryonic rat neurones.

The embryonic appearance of GABAergic cells and signals in the rat nervous system coincides with the appearance of transcripts encoding some but not all of the subunits forming GABA(A) receptor/Cl- channels. Quantitative in situ hybridization studies reveal higher variabilities in alpha2 and alpha3 subunit transcripts relative to others examined (alpha5, beta2, beta3 and gamma2) in six spinal and supraspinal regions. Immunocytochemistry of cells dissociated from the embryonic CNS shows that alpha2 and alpha3 subunits are detectable in differentiating neurones. FACS analyses of dissociated cells immunostained with alpha2- or alpha3- antibodies reveal immunopositive subpopulations of variable size in each region. Whole-cell recordings of acutely adherent neurones show that GABA activates Cl- currents whose fluctuations characteristically vary depending on a neurone's region of origin. Spectral analyses indicate a predominance of the low frequency (< 5 Hz) components, which vary regionally. Regression analyses reveal that (i) channel properties correlate with subunit transcript levels and (ii) dominant channel kinetics correlate with alpha2 and alpha3 subunit transcripts indexed as a ratio and with coexpressions of alpha5 and beta3. The correlations strongly suggest that alpha3 subunits in embryonic neurones are expressed in native receptor/channel complexes with slower kinetics than those containing alpha2 without alpha3 subunits. Thus, GABA(A) receptor/Cl- channels in these embryonic neurones may be encoded by the six transcripts (alpha2, alpha3, alpha5, and beta2, beta3, and gamma2) with proportions of alpha2, alpha3, alpha5, and beta3 subunits critical in determining their dominant kinetics.

Animals↗

PubMed1998

Initially expressed early rat embryonic GABA(A) receptor Cl- ion channels exhibit heterogeneous channel properties.

We have studied the earliest expression of GABA-induced CI- channels in the rat embryonic dorsal spinal cord (DSC) using in situ hybridization, immunocytochemistry, flow cytometry and electrophysiology. At embryonic day 13 (E13) cells in the dorsal region are still proliferating. In situ hybridization consistently showed transcripts encoding only three GABAA receptor subunits (alpha4, beta1 and gammal); immunocytochemistry both in tissue sections and in acutely isolated cells in suspension demonstrated the expression of the corresponding proteins and also revealed staining for other subunits (alpha2, alpha3, beta3, gamma2). In patch-recordings performed in cells acutely isolated from the dorsal cord, responses to GABA were detected in 356 out of 889 cells. GABA-evoked responses, which often displayed the opening of a few channels, were mediated by CI- ions, were inhibited by bicuculline and picrotoxin, and potentiated by benzodiazepines. Taken together, these observations indicate that CI- channels likely involve GABAA type receptors. Fluctuation analysis revealed channel kinetics consisting of three exponential components (Ts: approximately 1,9 and 90 ms) and a wide variety of inferred unitary conductance values, ranging between 4 and 40 pS. A comparison of these results with observations in other, later embryonic cell types and recombinant receptors suggests that most of the earliest E13 DSC GABAA receptors may include alpha3 subunit. These GABAA receptor Cl- channels may be activated physiologically as both GABA synthesizing enzymes and GABA are present in the E13 dorsal cord.

Animals↗

PubMed1998

Increase of preprotachykinin mRNA and substance P immunoreactivity in spared dorsal root ganglion neurons following partial sciatic nerve injury.

Complete sciatic nerve injury reduces substance P (SP) expression in primary sensory neurons of the L4 and L5 dorsal root ganglia (DRG), due to loss of target-derived nerve growth factor (NGF). Partial nerve injury spares a proportion of DRG neurons, whose axons lie in the partially degenerating nerve, and are exposed to elevated NGF levels from Schwann and other endoneurial cells involved in Wallerian degeneration. To test the hypothesis that SP is elevated in spared DRG neurons following partial nerve injury, we compared the effects of complete sciatic nerve transection (CSNT) with those of two types of partial injury, partial sciatic nerve transection (PSNT) and chronic constriction injury (CCI). As expected, a CSNT profoundly decreased SP expression at 4 and 14 days postinjury, but after PSNT and CCI the levels of preprotachykinin (PPT) mRNA, assessed by in situ hybridization, and the SP immunoreactivity (SP-IR) of the L4 and L5 DRGs did not decrease, nor did dorsal horn SP-IR decrease. Using retrograde labelling with fluorogold to identify spared DRG neurons, we found that the proportion of these neurons expressing SP-IR 14 days after injury was much higher than in neurons of normal DRGs. Further, the highest levels of SP-IR in individual neurons were detected in ipsilateral L4 and L5 DRG neurons after PSNT and CCI. We conclude that partial sciatic nerve injury elevates SP levels in spared DRG neurons. This phenomenon might be involved in the development of neuropathic pain, which commonly follows partial nerve injury.

Animals↗

PubMed1998

Inhibitory activity of boswellic acids from Boswellia serrata against human leukemia HL-60 cells in culture.

Four major triterpene acids including beta-boswellic acid (1), 3-O-acetyl-beta-boswellic acid (2), 11-keto-beta-boswellic acid (3), and 3-O-acetyl-11-keto-beta-boswellic acid (4) were isolated from the gum resin of Boswellia serrata and examined for their in vitro antitumor activity. They inhibited the synthesis of DNA, RNA and protein in human leukemia HL-60 cells in a dose dependent manner with IC50 values ranging from 0.6 to 7.1 microM. Among them, compound 4 induced the most pronounced inhibitory effects on DNA, RNA and protein synthesis with IC50 values of 0.6, 0.5, and 4.1 microM, respectively. The effect of 4 on DNA synthesis was found to be irreversible. Compound 4 significantly inhibited the cellular growth of HL-60 cells, but did not affect cell viability.

Antineoplastic Agents, Phytogenic↗

PubMed1998

Specific antibodies to the external vestibule of voltage-gated potassium channels block current.

Using delayed-rectifier potassium channels as examples, we have designed two specific blockers by generating specific antipeptide antibodies to epitopes in the external vestibules of two channel proteins, Kv1.2 and Kv3.1. These antibodies reduced whole-cell Kv1.2 or Kv3.1 currents in transfected cells and the effect was blocked by the corresponding peptide antigen, but not by control peptides. A control antibody had little effect on Kv1.2 currents and the Kv1.2 blocker antibody had limited effect on other related potassium currents. Furthermore, the Kv1.2 blocking antibody inhibited dendrotoxin binding to Kv1.2 channel proteins in transfected cells. Moreover, using the Kv1.2 blocker antibody, we determined the presence and relative contribution of endogenous Kv1.2 to the overall endogenous K+ currents in NG108 neuronal cells. This guided design of specific channel blockers will facilitate future physiological studies on ion channel functions.

Amino Acid Sequence↗