Keith Bonin

Light torque nanocontrol, nanomotors and nanorockers.

In a novel application of light torques, we manipulate and control the rotation of nanorods. We apply light torques to 250 nm diameter glass nanorods in a single-beam optical trap. Light-torque operated nanomotors whir at moderate speeds that depend on several factors, including the magnitude of the light torque, the viscosity of the surrounding medium, and the rotation rate of the electric field vector of the linearly polarized trapping light. Two new modes of behavior - rocking motion and saltatory motion - are also described and explained.

Aversive stimulus differentially triggers subsecond dopamine release in reward regions

Aversive stimuli have a powerful impact on behavior and are considered to be the opposite valence of pleasure. Recent studies have determined some populations of ventral tegmental area (VTA) dopaminergic neurons are activated by several types of aversive stimuli, whereas other distinct populations are either inhibited or unresponsive. However, it is not clear where these aversion-responsive neurons project, and whether alterations in their activity translate into dopamine release in the terminal field. Here we show unequivocally that the neurochemical and anatomical substrates responsible for the perception and processing of pleasurable stimuli within the striatum are also activated by tail pinch, a classical painful and aversive stimulus. Dopamine release is triggered in the dorsal striatum and nucleus accumbens (NAc) core by tail pinch and is time locked to the duration of the stimulus, indicating that the dorsal striatum and NAc core are neural substrates, which are involved in the perception of aversive stimuli. However, dopamine is released in the NAc shell only when tail pinch is removed, indicating that the alleviation of aversive condition could be perceived as a rewarding event.

Optogenetic control of striatal dopamine release in rats

J. Neurochem. (2010) 114, 1344–1352.

Absolute measurement of the optical polarizability of C60

We report on a new optical technique that uses light forces and a time-of-flight spectrometer to make absolute measurements of cluster polarizabilities. This is also the first accurate report of an ac polarizability measurement of a condensable cluster in the gas phase. We have determined the optical polarizability of C60 at the fundamental wavelength of a Nd:YAG laser (1.064 μm) to be α=79±4 A3.

Determining the mechanical properties of electrospun poly-ε-caprolactone (PCL) nanofibers using AFM and a novel fiber anchoring technique

Due to its low cost, biocompatibility and slow bioresorption, poly-ε-caprolactone (PCL) continues to be a suitable material for select biomedical engineering applications. We used a combined atomic force microscopy (AFM)/optical microscopy technique to determine key mechanical properties of individual electrospun PCL nanofibers with diameters between 440-1040nm. Compared to protein nanofibers, PCL nanofibers showed much lower adhesion, as they slipped on the substrate when mechanically manipulated. We, therefore, first developed a novel technique to anchor individual PCL nanofibers to micrometer-sized ridges on a substrate, and then mechanically tested anchored nanofibers. When held at constant strain, tensile stress relaxed with fast and slow relaxation times of 1.0±0.3s and 8.8±3.1s, respectively. The total tensile modulus was 62±26MPa, the elastic (non-relaxing) component of the tensile modulus was 53±36MPa. Individual PCL fibers could be stretched elastically (without permanent deformation) to strains of 19-23%. PCL nanofibers are rather extensible; they could be stretched to a strain of at least 98%, and a tensile strength of at least 12MPa, before they slipped off the AFM tip. PCL nanofibers that had aged for over a month at ambient conditions became stiffer and less elastic. Our technique provides accurate nanofiber mechanical data, which are needed to guide construction of scaffolds for cells and other biomedical devices.

Easy and direct method for calibrating atomic force microscopy lateral force measurements

We have designed and tested a new, inexpensive, easy-to-make and easy-to-use calibration standard for atomic force microscopy (AFM) lateral force measurements. This new standard simply consists of a small glass fiber of known dimensions and Youngs modulus, which is fixed at one end to a substrate and which can be bent laterally with the AFM tip at the other end. This standard has equal or less error than the commonly used method of using beam mechanics to determine a cantilevers lateral force constant. It is transferable, thus providing a universal tool for comparing the calibrations of different instruments. It does not require knowledge of the cantilever dimensions and composition or its tip height. This standard also allows direct conversion of the photodiode signal to force and, thus, circumvents the requirement for a sensor response (sensitivity) measurement.

Characterizing the micro-scale elastic modulus of hydrogels for use in regenerative medicine.

Our objective was to characterize the elasticity of hydrogel formulations intended to mimic physical properties that cells and tissues experience in vivo. Using atomic force microscopy (AFM), we tested a variety of concentrations in a variety of biomaterials, including agarose, alginate, the collagens, fibrin, hyaluronic acid, kerateine, laminin, Matrigel, polyacrylamide, polyethylene glycol diacrylate (PEGDA) and silicone elastomer (polydimethylsiloxane). Manipulations of the concentration of biomaterials were detectable in AFM measurements of elasticity (Youngs modulus, E), and E tended to increase with increased concentration. Depending on the biomaterials chosen, and their concentrations, generation of tunable biocompatible hydrogels in the physiologic range is possible.

Optogenetic stimulation of VTA dopamine neurons reveals that tonic but not phasic patterns of dopamine transmission reduce ethanol self-administration.

There is compelling evidence that acute ethanol exposure stimulates ventral tegmental area (VTA) dopamine cell activity and that VTA-dependent dopamine release in terminal fields within the nucleus accumbens plays an integral role in the regulation of ethanol drinking behaviors. Unfortunately, due to technical limitations, the specific temporal dynamics linking VTA dopamine cell activation and ethanol self-administration are not known. In fact, establishing a causal link between specific patterns of dopamine transmission and ethanol drinking behaviors has proven elusive. Here, we sought to address these gaps in our knowledge using a newly developed viral-mediated gene delivery strategy to selectively express Channelrhodopsin-2 (ChR2) on dopamine cells in the VTA of wild-type rats. We then used this approach to precisely control VTA dopamine transmission during voluntary ethanol drinking sessions. The results confirmed that ChR2 was selectively expressed on VTA dopamine cells and delivery of blue light pulses to the VTA induced dopamine release in accumbal terminal fields with very high temporal and spatial precision. Brief high frequency VTA stimulation induced phasic patterns of dopamine release in the nucleus accumbens. Lower frequency stimulation, applied for longer periods mimicked tonic increases in accumbal dopamine. Notably, using this optogenetic approach in rats engaged in an intermittent ethanol drinking procedure, we found that tonic, but not phasic, stimulation of VTA dopamine cells selectively attenuated ethanol drinking behaviors. Collectively, these data demonstrate the effectiveness of a novel viral targeting strategy that can be used to restrict opsin expression to dopamine cells in standard outbred animals and provide the first causal evidence demonstrating that tonic activation of VTA dopamine neurons selectively decreases ethanol self-administration behaviors.

Dopamine Uptake Changes Associated with Cocaine Self-Administration

The present study was designed to reveal the relationship between cocaine-induced dopamine uptake changes and patterns of cocaine self-administration observed under a fixed-ratio schedule. Cocaine was intravenously infused into anesthetized rats, according to inter-infusion intervals obtained from self-administering animals, and dopamine uptake changes (apparent Km) were assessed in the nucleus accumbens using voltammetry. The data demonstrate that cocaine-induced dopamine transporter (DAT) inhibition accounts for the accumbal dopamine fluctuations, which are associated with the cyclic regularity of cocaine intake observed during self-administration. Specifically, the inter-infusion intervals that are maintained during cocaine self-administration correlate with the maintenance of a rapidly changing level of dopamine uptake inhibition, which appears to be tightly regulated. Furthermore, this maintained level of dopamine uptake inhibition was found to shift upward using intervals from animals that had shown an escalation in the rate of cocaine self-administration. Although no significant change in the apparent Km was revealed in animals that exhibited an escalation in the rate of cocaine intake, an increased dopamine uptake rate was found suggesting an upregulation of DAT number in response to a history of high cocaine intake. This is the first demonstration of the tight correlation that exists between the level of dopamine uptake inhibition and rates of cocaine self-administration. Moreover, a new mathematical model was created that quantitatively describes the changes in cocaine-induced dopamine uptake and correctly predicts the level of dopamine uptake inhibition. This model permits a computational interpretation of cocaine-induced dopamine uptake changes during cocaine self-administration.

Forces Required of Kinesin during Processive Transport through Cytoplasm

The purpose of this paper is to deduce whether the maximum force, steplike movement, and rate of ATP consumption of kinesin, as measured in buffer, are sufficient for the task of fast transport of vesicles in cells. Our results show that moving a 200-nm vesicle in viscoelastic COS7 cytoplasm, with the same steps as observed for kinesin-driven beads in buffer, required a maximum force of 16 pN and work per step of 1 +/- 0.7 ATP, if the drag force was assumed to decrease to zero between steps. In buffer, kinesin can develop a force of 6-7 pN while consuming 1 ATP/step, comparable to the required values. As an alternative to assuming that the force vanishes between steps, the measured COS7 viscoelasticity was extrapolated to zero frequency by a numerical fit. The force required to move the bead then exceeded 75 pN at all times and peaked briefly to 92 pN, well beyond the measured capabilities of a single kinesin in buffer. The work per step increased to 7 +/- 5 ATP, greatly exceeding the energy available to a single motor.