Electrostatic Contributions of Aromatic Residues in the Local Anesthetic Receptor of Voltage-Gated Sodium Channels

Antiarrhythmics, anticonvulsants, and local anesthetics target voltage-gated sodium channels, decreasing excitability of nerve and muscle cells. Channel inhibition by members of this family of cationic, hydrophobic drugs relies on the presence of highly conserved aromatic residues in the pore-lining S6 segment of the fourth homologous domain of the channel. We tested whether channel inhibition was facilitated by an electrostatic attraction between lidocaine and {pi} electrons of the aromatic rings of these residues, namely a cation-{pi} interaction. To this end, we used the in vivo nonsense suppression method to incorporate a series of unnatural phenylalanine derivatives designed to systematically reduce the negative electrostatic potential on the face of the aromatic ring. In contrast to standard point mutations at the same sites, these subtly altered amino acids preserve the wild-type voltage dependence of channel activation and inactivation. Although these phenylalanine derivatives have no effect on low-affinity tonic inhibition by lidocaine or its permanently charged derivative QX-314 at any of the substituted sites, high-affinity use-dependent inhibition displays substantial cation-{pi} energetics for 1 residue only: Phe1579 in rNaV1.4. Replacement of the aromatic ring of Phe1579 by cyclohexane, for example, strongly reduces use-dependent inhibition and speeds recovery of lidocaine-engaged channels. Channel block by the neutral local anesthetic benzocaine is unaffected by the distribution of {pi} electrons at Phe1579, indicating that our aromatic manipulations expose electrostatic contributions to channel inhibition. These results fine tune our understanding of local anesthetic inhibition of voltage-gated sodium channels and will help the design of safer and more salutary therapeutic agents

Mergers, Migration, and Signaling

The interaction between learning at the individual level and the trajectory of a population over time is fundamental to our understanding of linguistic change. Here we use game theory as a mathematical framework for formulating and testing hypotheses about this interaction. We formalize two hypotheses regarding the spread of vowel mergers and use them to derive the proportion of merged in-migrants that would precipitate a merger in a previously non-merged speech community. We test these predictions against the documented spread of the low-back merger. In light of these results, we consider the impact of social network structures on both models

I am a Monster: An Exploration of the Self through Examination of Fragmented Identity or Mary Shelley’s Frankenstein Becomes a Guide for Self-Reflection

The purpose of this thesis was to explore the ways a fragmented identity can be reconciled through examination and analysis of Mary Shelley’s Frankenstein and several other works of art. Findings suggest that identity is both generated by and projected onto individuals, and reconciliation of these questions can turn the concept of monstrosity from a negative to a positive. This research supports and promotes the notion that individuals are more than simply the sum of all their parts, and that identities can simultaneously endure the paradox of being fragmented yet whole

A Cation–π Interaction between Extracellular TEA and an Aromatic Residue in Potassium Channels

Open-channel blockers such as tetraethylammonium (TEA) have a long history as probes of the permeation pathway of ion channels. High affinity blockade by extracellular TEA requires the presence of an aromatic amino acid at a position that sits at the external entrance of the permeation pathway (residue 449 in the eukaryotic voltage-gated potassium channel Shaker). We investigated whether a cation–{pi} interaction between TEA and such an aromatic residue contributes to TEA block using the in vivo nonsense suppression method to incorporate a series of increasingly fluorinated Phe side chains at position 449. Fluorination, which is known to decrease the cation–{pi} binding ability of an aromatic ring, progressively increased the inhibitory constant Ki for the TEA block of Shaker. A larger increase in Ki was observed when the benzene ring of Phe449 was substituted by nonaromatic cyclohexane. These results support a strong cation–{pi} component to the TEA block. The data provide an empirical basis for choosing between Shaker models that are based on two classes of reported crystal structures for the bacterial channel KcsA, showing residue Tyr82 in orientations either compatible or incompatible with a cation–{pi} mechanism. We propose that the aromatic residue at this position in Shaker is favorably oriented for a cation–{pi} interaction with the permeation pathway. This choice is supported by high level ab initio calculations of the predicted effects of Phe modifications on TEA binding energy

Alien Registration- Ahern, Mary A. (Portland, Cumberland County)

https://digitalmaine.com/alien_docs/22142/thumbnail.jp

A Cation-π Interaction Discriminates among Sodium Channels That Are Either Sensitive or Resistant to Tetrodotoxin Block

Voltage-gated sodium channels control the upstroke of the action potential in excitable cells of nerve and muscle tissue, making them ideal targets for exogenous toxins that aim to squelch electrical excitability. One such toxin, tetrodotoxin (TTX), blocks sodium channels with nanomolar affinity only when an aromatic Phe or Tyr residue is present at a specific location in the external vestibule of the ion-conducting pore. To test whether TTX is attracted to Tyr401 of NaV1.4 through a cation-{pi} interaction, this aromatic residue was replaced with fluorinated derivatives of Phe using in vivo nonsense suppression. Consistent with a cation-{pi} interaction, increased fluorination of Phe401, which reduces the negative electrostatic potential on the aromatic face, caused a monotonic increase in the inhibitory constant for block. Trifluorination of the aromatic ring decreased TTX affinity by ~50-fold, a reduction similar to that caused by replacement with the comparably hydrophobic residue Leu. Furthermore, we show that an energetically equivalent cation-{pi} interaction underlies both use-dependent and tonic block by TTX. Our results are supported by high level ab initio quantum mechanical calculations applied to a model of TTX binding to benzene. Our analysis suggests that the aromatic side chain faces the permeation pathway where it orients TTX optimally and interacts with permeant ions. These results are the first of their kind to show the incorporation of unnatural amino acids into a voltage-gated sodium channel and demonstrate that a cation-{pi} interaction is responsible for the obligate nature of an aromatic at this position in TTX-sensitive sodium channels

How Uniqueness Guides Definite Description Processing

Most analyses of definiteness are based on two important notions: uniqueness and familiarity. Fundamentally, both approaches ascribe some content to the conventional meaning of definite, but not indefinite, descriptions. We explore the effect of determiner choice on listeners’ expectations about possible referents using eye-tracking in the visual world paradigm. We present listeners with temporarily ambiguous definite descriptions where a single referent is unique under the greatest number of possible semantic descriptions. We find that uniqueness is not only a robust notion for describing the meaning of definitess, but also a crucial factor in guiding listeners in the online processing of definite descriptions

How Uniqueness Guides Definite Description Processing

Most analyses of definiteness are based on two important notions: uniqueness and familiarity. Fundamentally, both approaches ascribe some content to the conventional meaning of definite, but not indefinite, descriptions. We explore the effect of determiner choice on listeners’ expectations about possible referents using eye-tracking in the visual world paradigm. We present listeners with temporarily ambiguous definite descriptions where a single referent is unique under the greatest number of possible semantic descriptions. We find that uniqueness is not only a robust notion for describing the meaning of definitess, but also a crucial factor in guiding listeners in the online processing of definite descriptions