A Ligand Channel through the G Protein Coupled Receptor Opsin

The G protein coupled receptor rhodopsin contains a pocket within its seven-transmembrane helix (TM) structure, which bears the inactivating 11-cis-retinal bound by a protonated Schiff-base to Lys296 in TM7. Light-induced 11-cis-/all-trans-isomerization leads to the Schiff-base deprotonated active Meta II intermediate. With Meta II decay, the Schiff-base bond is hydrolyzed, all-trans-retinal is released from the pocket, and the apoprotein opsin reloaded with new 11-cis-retinal. The crystal structure of opsin in its active Ops* conformation provides the basis for computational modeling of retinal release and uptake. The ligand-free 7TM bundle of opsin opens into the hydrophobic membrane layer through openings A (between TM1 and 7), and B (between TM5 and 6), respectively. Using skeleton search and molecular docking, we find a continuous channel through the protein that connects these two openings and comprises in its central part the retinal binding pocket. The channel traverses the receptor over a distance of ca. 70 Å and is between 11.6 and 3.2 Å wide. Both openings are lined with aromatic residues, while the central part is highly polar. Four constrictions within the channel are so narrow that they must stretch to allow passage of the retinal β-ionone-ring. Constrictions are at openings A and B, respectively, and at Trp265 and Lys296 within the retinal pocket. The lysine enforces a 90° elbow-like kink in the channel which limits retinal passage. With a favorable Lys side chain conformation, 11-cis-retinal can take the turn, whereas passage of the all-trans isomer would require more global conformational changes. We discuss possible scenarios for the uptake of 11-cis- and release of all-trans-retinal. If the uptake gate of 11-cis-retinal is assigned to opening B, all-trans is likely to leave through the same gate. The unidirectional passage proposed previously requires uptake of 11-cis-retinal through A and release of photolyzed all-trans-retinal through B

Atypical Antipsychotic Haloperidol Disrupts Prepulse Inhibition of Acoustic Startle Reflex in Larval Zebrafish

Prepulse inhibition (PPI) in larval zebrafish can be utilized as a behavioral marker for neural dysfunction. In this study, we show that application of pharmacological agents to affect the Mauthner Cell mediated startle movement in response to acoustic stimuli can be used to quantify dysfunction. By establishing a robust protocol, we were able to confirm the effects of numerous pharmacological agents on PPI. We were also able to establish that haloperidol had a negative effect on PPI. This parallels some recent mammalian PPI studies that indicate that haloperidol has mixed effect on PPI. Dose dependent effects, timing, and even the nature of the reductionist zebrafish model may be responsible for this difference. However, the large sample sizes and simple neural circuitry makes the larval zebrafish and excellent model for high throughput screens of psychotropic therapeutic agents

Student Satisfaction and Learning Outcomes in Asynchronous Online Lecture Videos.

Our study identified online lecture video styles that improved student engagement and satisfaction, while maintaining high learning outcomes in online education. We presented different lecture video styles with standardized material to students and then measured learning outcomes and satisfaction with a survey and summative assessment. We created an iterative qualitative coding scheme, coding online asynchronous lectures (COAL), to analyze open-ended student survey responses. Our results reveal that multimedia learning can be satisfying and effective. Students have strong preferences for certain video styles despite their equal learning outcomes, with the Learning Glass style receiving the highest satisfaction ratings. Video styles that were described as impersonal and unfamiliar were rated poorly, while those that were described as personal and engaging and evoked positive affective responses were rated highly. The students in our study rated lecture video styles that aligned with Mayers multimedia learning principles as highly satisfying, indicating that student feedback can be a valuable resource for course designers to consider as they design their own online courses. Finally, we provide guidelines for creating engaging, effective, and satisfying asynchronous lecture videos to support establishment of best practices in online instruction

Structural features of the opsin ligand channel.

<p>Coplanar cut through opsin revealing the channel with opening A, B and constrictions C1-C4 (a, top view). The position of Lys296 is indicated by a yellow dot. All-<i>trans</i>-retinal (green) is docked into the binding pocket (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#s4" target="_blank">Methods</a>). Electrostatic surface potentials were calculated using the program APBS <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#pone.0004382-Baker1" target="_blank">[22]</a> with nonlinear Poisson-Boltzmann equation and contoured at ±20kT/e and negatively and positively charged surface areas in red and blue, respectively (at high kT/e values the contour level is shifted from coloured to grey scale). (b, c) Side-views, with electrostatic surface potentials contoured at ±8kT/e. (d, e) Close-ups of openings A and B, defined by the residues as indicated.</p

Docking of retinal isomers to docking site I.

<p>Flexible docking of (a–c) 11<i>-cis-</i>retinal and (d–f) all-<i>trans</i>-retinal to site I located between opening A and C2 at the 90° kink of the channel (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#pone-0004382-g004" target="_blank">Figure 4</a>). The crystal structure of Ops* (PDB entry 3CAP) was used and full flexibility for Lys296 side chain was allowed. The most likely conformation of (a) 11<i>-cis-</i>retinal and (d) all-<i>trans</i>-retinal is shown together with the neighbouring residues. The conformation of Lys296 obtained by the docking procedure (orange) is superimposed to the starting conformation (light green). Cluster of docking poses of (b) 11<i>-cis-</i>retinal and (e) all-<i>trans</i>-retinal and (c, f) the respective lists of ranked docking poses at different RMSD cut-off values (different colours identify individual poses). The best scored pose of the finally selected cluster (shaded) is shown with ball and sticks in a, b, d and e.</p

Docking of retinal isomers to docking site II.

<p>Flexible docking of (a–c) 11<i>-cis-</i>retinal and (d–f) all-<i>trans</i>-retinal to site II, i.e. the retinal binding pocket (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#pone-0004382-g004" target="_blank">Figure 4</a>). The crystal structure of Ops* (PDB entry 3CAP) was used and full flexibility for Tyr191, Val204, Phe208, Phe273 and Lys296 side chains was allowed. The most likely conformation of (a) 11<i>-cis-</i>retinal and (d) all-<i>trans</i>-retinal is shown together with the neighbouring residues. The residues with altered conformation (orange) are super­imposed to the starting conformation. Cluster of docking poses of (b) 11<i>-cis-</i>retinal and (e) all-<i>trans</i>-retinal and (c, f) the respective list of ranked docking poses at different RMSD cut-off values (different colours identify individual poses). The best scored pose of the finally selected cluster (shaded) is shown with ball and sticks in a, b, d and e.</p

Location of the retinal docking sites.

<p>(a) The docking sites (I, II, III) are restricted to all residues within the radius of 10 Å (circles) from Met44 (I), Tyr268 (II) and Ala269 (III), respectively. Site I is close to opening A and the 90° kink of the channel, site II represents the retinal binding pocket, and site III is close to opening B. (b) Selected final conformations of retinal isomers resulting from the three docking procedures (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#pone-0004382-g005" target="_blank">Figures 5</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#pone-0004382-g006" target="_blank"></a><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#pone-0004382-g007" target="_blank">7</a> for details). Blue, 11-<i>cis-</i>retinal docked to site I; green, all-<i>trans</i>-retinal docked to site II; and cyan, all-<i>trans</i>-retinal docked to site III. (c) View onto the ligand channel (electrostatic surface potentials as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004382#pone-0004382-g001" target="_blank">Figure 1a</a>) with docked 11-<i>cis</i>- (blue) and all-<i>trans</i>-retinal (green, cyan). Parts of the receptor were omitted to visualize the two openings (A and B), the constrictions (C1–C4) and the neighbouring cavity (NC).</p

Rapid habituation of a touch-induced escape response in Zebrafish (Danio rerio) Larvae.

Zebrafish larvae have several biological features that make them useful for cellular investigations of the mechanisms underlying learning and memory. Of particular interest in this regard is a rapid escape, or startle, reflex possessed by zebrafish larvae; this reflex, the C-start, is mediated by a relatively simple neuronal circuit and exhibits habituation, a non-associative form of learning. Here we demonstrate a rapid form of habituation of the C-start to touch that resembles the previously reported rapid habituation induced by auditory or vibrational stimuli. We also show that touch-induced habituation exhibits input specificity. This work sets the stage for in vivo optical investigations of the cellular sites of plasticity that mediate habituation of the C-start in the larval zebrafish

Constriction sites within the channel.

<p>Minimum inner width (d_min) measured at intervals of 0.6 Å progressing from opening A to opening B. The residues defining the constriction sites (C1–C4) are indicated, as well as the maximum and minimum extensions of the β-ionone moiety of the retinal.</p