Molecular ontology of the parabrachial nucleus.

Diverse neurons in the parabrachial nucleus (PB) communicate with widespread brain regions. Despite evidence linking them to a variety of homeostatic functions, it remains difficult to determine which PB neurons influence which functions because their subpopulations intermingle extensively. An improved framework for identifying these intermingled subpopulations would help advance our understanding of neural circuit functions linked to this region.

Combined magnetic resonance and fluorescence imaging of the living mouse brain reveals glioma response to chemotherapy.

Fluorescent molecular tomographic (FMT) imaging can noninvasively monitor molecular function in living animals using specific fluorescent probes. However, macroscopic imaging methods such as FMT generally exhibit low anatomical details. To overcome this, we report a quantitative technique to image both structure and function by combining FMT and magnetic resonance (MR) imaging.

Rapid and modifiable neurotransmitter receptor dynamics at a neuronal synapse in vivo.

Synaptic plasticity underlies the adaptability of the mammalian brain, but has been difficult to study in living animals. Here we imaged the synapses between pre- and postganglionic neurons in the mouse submandibular ganglion in vivo, focusing on the mechanisms that maintain and regulate neurotransmitter receptor density at postsynaptic sites. Normally, synaptic receptor densities were maintained by rapid exchange of receptors with nonsynaptic regions (over minutes) and by continual turnover of cell surface receptors (over hours).

In vivo imaging of presynaptic terminals and postsynaptic sites in the mouse submandibular ganglion.

Much of what is currently known about the behavior of synapses in vivo has been learned at the mammalian neuromuscular junction, because it is large and accessible and also its postsynaptic acetylcholine receptors (AChRs) are readily labeled with a specific, high-affinity probe, alpha-bungarotoxin (BTX). Neuron-neuron synapses have thus far been much less accessible. We therefore developed techniques for imaging interneuronal synapses in an accessible ganglion in the peripheral nervous system.

Rapid synapse elimination after postsynaptic protein synthesis inhibition in vivo.

To examine the role of retrograde signals on synaptic maintenance, we inhibited protein synthesis in individual postsynaptic cells in vivo while monitoring presynaptic terminals. Within 12 h, axon terminals begin to atrophy and withdraw from normal postsynaptic sites. Structural similarities between this process and naturally occurring synapse elimination suggest that short-lived target derived factors not only participate in synaptic maintenance in adults, but also regulate elimination of connections during development.

The cholinergic antagonist alpha-bungarotoxin also binds and blocks a subset of GABA receptors.

The polypeptide snake toxin alpha-bungarotoxin (BTX) has been used in hundreds of studies on the structure, function, and development of the neuromuscular junction because it binds tightly and specifically to the nicotinic acetylcholine receptors (nAChRs) at this synapse. We show here that BTX also binds to and blocks a subset of GABA(A) receptors (GABA(A)Rs) that contain the GABA(A)R beta3 subunit. These results introduce a previously unrecognized tool for analysis of GABA(A)Rs but may complicate interpretation of some studies on neuronal nAChRs.

Peptide tags for labeling membrane proteins in live cells with multiple fluorophores.

We describe a method to label specific membrane proteins with fluorophores for live imaging. Fusion proteins are generated that incorporate into their extracellular domains short peptide sequences (13-38 amino acids) recognized with high affinity and specificity by protein ligands, alpha-bungarotoxin (BTX), or streptavidin (SA). Many fluorophore- and enzyme-conjugated derivatives of both ligands are commercially available.