Regional and sex differences in spontaneous striatal dopamine transmission.

Striatal dopamine release is key for learning and motivation and is composed of subregions including the dorsal striatum (DS), nucleus accumbens core, and the nucleus accumbens shell. Spontaneously occurring dopamine release was compared across these subregions. Dopamine release/uptake dynamics differ across striatal subregions, with dopamine transient release amplitude and release frequency greatest in male mice, and the largest signals observed in the DS. Surprisingly, female mice exhibited little regional differences in dopamine release for DS and nucleus accumbens core regions, but lower release in the nucleus accumbens shell. Blocking voltage-gated K+ channel (Kv channels) with 4-aminopyridine enhanced dopamine detection without affecting reuptake. The 4-aminopyridine effects were greatest in ventral regions of female mice, suggesting regional differences in Kv channel expression. The dopamine transporter blocker cocaine also enhanced detection across subregions in both sexes, with greater overall increased release in females than males. Thus, sex differences in dopamine transmission are apparent and likely include differences in the Kv channel and dopamine transporter function. The lack of regional differences in dopamine release observed in females indicates differential regulation of spontaneous and evoked dopamine release.

Application of a Skin Adhesive to Maintain Seal in Negative Pressure Wound Therapy: Demonstration of a New Technique.

Optimal wound healing with negative pressure wound therapy (NPWT) relies on a properly sealed vacuum system. Anatomically difficult wounds impair the adhesive dressing, which results in air leaks that disrupt the integrity of the NPWT system and hinder wound healing. OBJECTIVE: The authors demonstrate a new technique using a cyanoacrylate-based tissue adhesive to maintain an airtight, durable seal in NPWT. MATERIALS AND METHODS: A 52-year-old woman with a degloving injury to the right thigh extending into the groin, resulting in massive necrosis, presented to the emergency department. Using a skin closure system, 2 polyester mesh tape strips were placed near the perineal region of the wound to reinforce the adhesive drape of the NPWT system. Skin grafts were applied over the wound after about 3 weeks of NPWT, and the skin closure system was applied in the same fashion to reinforce the adhesive drape. RESULTS: An airtight seal was consistently maintained for several days in between dressing changes. The size of the wound was visibly reduced at each dressing change. An airtight seal was maintained for 5 days after placement of the skin grafts; after 5 days, the dressing was removed without difficulty and skin irritation. The skin grafts appeared healthy with adequate tissue take. CONCLUSIONS: Maintaining an airtight seal in NPWT is crucial to wound healing. Cyanoacrylate tissue adhesives appear to be a safe and viable option for creating a durable seal in NPWT for wounds in anatomically difficult locations.

Spatial characterization of Bessel-like beams for strong-field physics.

We present a compact, simple design for the generation and tuning of both the spot size and effective focal length of Bessel-like beams. In particular, this setup provides an important tool for the use of Bessel-like beams with high-power, femtosecond laser systems. Using a shallow angle axicon in conjunction with a spherical lens, we show that it is possible to focus Bessel-like modes to comparable focal spot sizes to sharp axicons while maintaining a long effective focal length. The resulting focal profiles are characterized in detail using an accurate high dynamic range imaging technique. Quantitatively, we introduce a metric (R0.8) which defines the spot-size containing 80% of the total energy. Our setup overcomes the typical compromise between long working distances and small spot sizes. This is particularly relevant for strong-field physics where most experiments must operate in vacuum.

The Application of Skin Adhesive to Maintain Seal in Negative Pressure Wound Therapy.

BACKGROUND: Optimal wound healing in negative pressure wound therapy (NPWT) depends on a properly sealed vacuum system. Anatomically difficult wounds disrupt the adhesive dressing, resulting in air leaks that impair the integrity of this system. Several techniques have been used in previous reports to prevent air leaks, including the addition of skin adhesives (eg, Skin-Prep [Smith and Nephew, St. Petersburg, FL] or compound tincture of benzoin), hydrocolloid dressings, silicone, and stoma paste. The purpose of this case report is to demonstrate the effectiveness of using a cyanoacrylate tissue adhesive, dermaFLEX (FLEXCon, Spencer, MA), in maintaining an airtight, durable seal in NPWT. MATERIALS AND METHODS: The authors present a patient with a difficult to manage anogenital wound where efforts to maintain an airtight seal in NPWT proved difficult. It was decided during the course of treatment to use the cyanoacrylate tissue adhesive to create an airtight, durable seal. The tissue adhesive was applied circumferentially to the skin surrounding the wound edge. After placement of vacuum-assisted closure foam over the wound, the adhesive dressing was applied with its edges overlapping the skin area where the tissue adhesive was applied. RESULTS: The size of the wound was visibly reduced at each dressing change. An airtight seal was consistently maintained for 3 days at a time, surviving the difficult environment of the wound and maximizing the life of each adhesive dressing. CONCLUSION: For wounds in anatomically challenging locations, the use of the tissue adhesive appears to be a safe and viable option in creating a durable seal in NPWT.

Antibody elicited against the gp41 N-heptad repeat (NHR) coiled-coil can neutralize HIV-1 with modest potency but non-neutralizing antibodies also bind to NHR mimetics.

Following CD4 receptor binding to the HIV-1 envelope spike (Env), the conserved N-heptad repeat (NHR) region of gp41 forms a coiled-coil that is a precursor to the fusion reaction. Although it has been a target of drug and vaccine design, there are few monoclonal antibody (mAb) tools with which to probe the antigenicity and immunogenicity specifically of the NHR coiled-coil. Here, we have rescued HIV-1-neutralizing anti-NHR mAbs from immune phage display libraries that were prepared (i) from b9 rabbits immunized with a previously described mimetic of the NHR coiled-coil, N35(CCG)-N13, and (ii) from an HIV-1 infected individual. We describe a rabbit single-chain Fv fragment (scFv), 8K8, and a human Fab, DN9, which specifically recognize NHR coiled-coils that are unoccupied by peptide corresponding to the C-heptad repeat or CHR region of gp41 (e.g. C34). The epitopes of 8K8 and DN9 were found to partially overlap with that of a previously described anti-NHR mAb, IgG D5; however, 8K8 and DN9 were much more specific than D5 for unoccupied NHR trimers. The mAbs, including a whole IgG 8K8 molecule, neutralized primary HIV-1 of clades B and C in a pseudotyped virus assay with comparable, albeit relatively modest potency. Finally, a human Fab T3 and a rabbit serum (both non-neutralizing) were able to block binding of D5 and 8K8 to a gp41 NHR mimetic, respectively, but not the neutralizing activity of these mAbs. We conclude from these results that NHR coiled-coil analogs of HIV-1 gp41 elicit many Abs during natural infection and through immunization, but that due to limited accessibility to the corresponding region on fusogenic gp41 few can neutralize. Caution is therefore required in targeting the NHR for vaccine design. Nevertheless, the mAb panel may be useful as tools for elucidating access restrictions to the NHR of gp41 and in designing potential improvements to mimetics of receptor-activated Env.

An affinity-enhanced neutralizing antibody against the membrane-proximal external region of human immunodeficiency virus type 1 gp41 recognizes an epitope between those of 2F5 and 4E10.

The membrane-proximal external region (MPER) of human immunodeficiency virus type 1 (HIV-1) gp41 bears the epitopes of two broadly neutralizing antibodies (Abs), 2F5 and 4E10, making it a target for vaccine design. A third Ab, Fab Z13, had previously been mapped to an epitope that overlaps those of 2F5 and 4E10 but only weakly neutralizes a limited set of primary isolates. Here, libraries of Fab Z13 variants displayed on phage were engineered and affinity selected against an MPER peptide and recombinant gp41. A high-affinity variant, designated Z13e1, was isolated and found to be approximately 100-fold improved over the parental Fab not only in binding affinity for the MPER antigens but also in neutralization potency against sensitive HIV-1. Alanine scanning of MPER residues 664 to 680 revealed that N671 and D674 are crucial for peptide recognition as well as for the neutralization of HIV-1 by Z13e1. Ab competition studies and truncation of MPER peptides indicate that Z13e1 binds with high affinity to an epitope between and overlapping with those of 2F5 and 4E10, with the minimal peptide epitope WASLWNWFDITN. Still, Z13e1 remained about an order of magnitude less potent than 4E10 against several isolates of pseudotyped HIV-1. The sum of our molecular analyses with Z13e1 suggests that the segment on the MPER of gp41 between the 2F5 and 4E10 epitopes is exposed on the functional envelope trimer but that access to the specific Z13e1 epitope within this segment is limited. Thus, the ability of MPER-bearing immunogens to elicit potent HIV-1-neutralizing Abs may depend in part on recapitulating the particular constraints that the functional envelope trimer imposes on the segment of the MPER to which Z13e1 binds.

Structure-function analysis of the epitope for 4E10, a broadly neutralizing human immunodeficiency virus type 1 antibody.

The human immunodeficiency virus type 1 (HIV-1) neutralizing antibody 4E10 binds to a linear, highly conserved epitope within the membrane-proximal external region of the HIV-1 envelope glycoprotein gp41. We have delineated the peptide epitope of the broadly neutralizing 4E10 antibody to gp41 residues 671 to 683, using peptides with different lengths encompassing the previously suggested core epitope (NWFDIT). Peptide binding to the 4E10 antibody was assessed by competition enzyme-linked immunosorbent assay, and the K(d) values of selected peptides were determined using surface plasmon resonance. An Ala scan of the epitope indicated that several residues, W672, F673, and T676, are essential (>1,000-fold decrease in binding upon replacement with alanine) for 4E10 recognition. In addition, five other residues, N671, D674, I675, W680, and L679, make significant contributions to 4E10 binding. In general, the Ala scan results agree well with the recently reported crystal structure of 4E10 in complex with a 13-mer peptide and with our circular dichroism analyses. Neutralization competition assays confirmed that the peptide NWFDITNWLWYIKKKK-NH(2) could effectively inhibit 4E10 neutralization. Finally, to limit the conformational flexibility of the peptides, helix-promoting 2-aminoisobutyric acid residues and helix-inducing tethers were incorporated. Several peptides have significantly improved affinity (>1,000-fold) over the starting peptide and, when used as immunogens, may be more likely to elicit 4E10-like neutralizing antibodies. Hence, this study represents the first stage toward iterative development of a vaccine based on the 4E10 epitope.