Ivo Vanzetta

In-vivo Optical Imaging of Cortical Architecture and Dynamics

A number of new imaging techniques are available to scientists to visualize the functioning brain directly, revealing unprecedented details. These imaging techniques have provided a new level of understanding of the principles underlying cortical development, organization and function. In this chapter we will focus on optical imaging in the living mammalian brain, using two complementary imaging techniques. The first technique is based on intrinsic signals. The second technique is based on voltage-sensitive dyes. Currently, these two optical imaging techniques offer the best spatial and temporal resolution, but also have inherent limitations. We shall provide a few examples of new findings obtained mostly in work done in our laboratory. The focus will be upon the understanding of methodological aspects which in turn should contribute to optimal use of these imaging techniques. General reviews describing earlier work done on simpler preparations have been published elsewhere (Cohen, 1973; Tasaki and Warashina, 1976; Waggoner and Grinvald, 1977; Waggoner, 1979; Salzberg, 1983; Grinvald, 1984; Grinvald et al., 1985; De Weer and Salzberg, 1986; Cohen and Lesher, 1986; Salzberg et al., 1986; Loew, 1987; Orbach, 1987; Blasdel, 1988, 1989; Grinvald et al., 1988; Kamino, 1991; Cinelli and Kauer, 1992; Frostig, 1994).

Compartment-resolved imaging of activity-dependent dynamics of cortical blood volume and oximetry.

Optical imaging, positron emission tomography, and functional magnetic resonance imaging (fMRI) all rely on vascular responses to image neuronal activity. Although these imaging techniques are used successfully for functional brain mapping, the detailed spatiotemporal dynamics of hemodynamic events in the various microvascular compartments have remained unknown. Here we used high-resolution optical imaging in area 18 of anesthetized cats to selectively explore sensory-evoked cerebral blood-volume (CBV) changes in the various cortical microvascular compartments. To avoid the confounding effects of hematocrit and oximetry changes, we developed and used a new fluorescent blood plasma tracer and combined these measurements with optical imaging of intrinsic signals at a near-isosbestic wavelength for hemoglobin (565 nm). The vascular response began at the arteriolar level, rapidly spreading toward capillaries and venules. Larger veins lagged behind. Capillaries exhibited clear blood-volume changes. Arterioles and arteries had the largest response, whereas the venous response was smallest. Information about compartment-specific oxygen tension dynamics was obtained in imaging sessions using 605 nm illumination, a wavelength known to reflect primarily oximetric changes, thus being more directly related to electrical activity than CBV changes. Those images were radically different: the response began at the parenchyma level, followed only later by the other microvascular compartments. These results have implications for the modeling of fMRI responses (e.g., the balloon model). Furthermore, functional maps obtained by imaging the capillary CBV response were similar but not identical to those obtained using the early oximetric signal, suggesting the presence of different regulatory mechanisms underlying these two hemodynamic processes.

Evidence and lack of evidence for the initial dip in the anesthetized rat: implications for human functional brain imaging.

a s p s a Two important contributions by Lindauer et al. and Jones et al. on pages 988–1001 and 1002–1015, respectively, in this issue of NeuroImage describe cortical hemodynamics in the rat as derived from imaging spectroscopy and an advanced theoretical model calculating photon pathlength in brain tissue. Surprisingly, the Sheffield group has found evidence for an early deoxygenation phase (“the initial dip”) but the Berlin group did not. Here we try to explain the above puzzle. This question is of primary interest because numerous events at the molecular level are involved in neurovascular, neurometabolic, and glial–metabolic coupling to changing electrical activity in neurons, in intracellular as well as and extracellular compartments. Understanding the mechanistic role of the various participants in hemodynamic coupling to electrical activity at the level of their intricate spatiotemporal interactions depends on the availability of a detailed experimental description of what really happens during the first second following the onset of electrical activity. Evaluation of proposed models likewise requires such an experimental description. We shall focus on the small initial dip observed (or not) in the rat and on the implications of these findings derived from optical imaging and imaging spectroscopy to future improvements of the spatial resolution of human f-MRI.

Non-invasive visualization of cortical columns by fMRI

Functional magnetic resonance imaging can now resolve individual cortical columns, which should provide insights into sensory perception and higher cognitive functions.

Coupling between neuronal activity and microcirculation: Implications for functional brain imaging

In the neocortex, neurons with similar response properties are often clustered together in column‐like structures, giving rise to what has become known as functional architecture—the mapping of various stimulus feature dimensions onto the cortical sheet. At least partially, we owe this finding to the availability of several functional brain imaging techniques, both post‐mortem and in‐vivo, which have become available over the last two generations, revolutionizing neuroscience by yielding information about the spatial organization of active neurons in the brain. Here, we focus on how our understanding of such functional architecture is linked to the development of those functional imaging methodologies, especially to those that image neuronal activity indirectly, through metabolic or haemodynamic signals, rather than directly through measurement of electrical activity. Some of those approaches allow exploring functional architecture at higher spatial resolution than others. In particular, optical imaging of intrinsic signals reaches the striking detail of È50 yum, and, together with other methodologies, it has allowed characterizing the metabolic and haemodynamic responses induced by sensory‐evoked neuronal activity. Here, we review those findings about the spatio‐temporal characteristics of neurovascular coupling and discuss their implications for functional brain imaging, including position emission tomography, and non‐invasive neuroimaging techniques, such as funtional magnetic resonance imaging, applicable also to the human brain.

Accurate spike estimation from noisy calcium signals for ultrafast three-dimensional imaging of large neuronal populations in vivo

Extracting neuronal spiking activity from large-scale two-photon recordings remains challenging, especially in mammals in vivo, where large noises often contaminate the signals. We propose a method, MLspike, which returns the most likely spike train underlying the measured calcium fluorescence. It relies on a physiological model including baseline fluctuations and distinct nonlinearities for synthetic and genetically encoded indicators. Model parameters can be either provided by the user or estimated from the data themselves. MLspike is computationally efficient thanks to its original discretization of probability representations; moreover, it can also return spike probabilities or samples. Benchmarked on extensive simulations and real data from seven different preparations, it outperformed state-of-the-art algorithms. Combined with the finding obtained from systematic data investigation (noise level, spiking rate and so on) that photonic noise is not necessarily the main limiting factor, our method allows spike extraction from large-scale recordings, as demonstrated on acousto-optical three-dimensional recordings of over 1,000 neurons in vivo.

Elliptically polarized light for depth resolved optical imaging

It is shown that using elliptically polarized light permits selecting well-defined subsurface volumes in a turbid medium. This suggests the possibility of probing biological tissues at specific depths. First, we present the method and preliminary results obtained on an Intralipid phantom. We next report on the method’s performance on a biological phantom (chicken breast) and, finally, on the exposed cortex of an anesthetized rat.

Motion clouds: model-based stimulus synthesis of natural-like random textures for the study of motion perception

Choosing an appropriate set of stimuli is essential to characterize the response of a sensory system to a particular functional dimension, such as the eye movement following the motion of a visual scene. Here, we describe a framework to generate random texture movies with controlled information content, i.e., Motion Clouds. These stimuli are defined using a generative model that is based on controlled experimental parametrization. We show that Motion Clouds correspond to dense mixing of localized moving gratings with random positions. Their global envelope is similar to natural-like stimulation with an approximate full-field translation corresponding to a retinal slip. We describe the construction of these stimuli mathematically and propose an open-source Python-based implementation. Examples of the use of this framework are shown. We also propose extensions to other modalities such as color vision, touch, and audition.

Voltage-Sensitive Dye Imaging of Neocortical Activity

Neural computations underlying sensory perception, cognition, and motor control are performed by populations of neurons at different anatomical and temporal scales. Few techniques are currently available for exploring the dynamics of local and large range populations. Voltage-sensitive dye imaging (VSDI), based on organic voltage probes, reveals neural population activity in areas ranging from a few tens of micrometers to a couple of centimeters, or two areas up to ~10 cm apart. VSDI provides a submillisecond temporal resolution and a spatial resolution of ~50 µm. The dye signal emphasizes subthreshold synaptic potentials. VSDI has been applied in the mouse, rat, gerbil, ferret, tree shrew, cat, and monkey cortices to explore the lateral spread of retinotopic or somatotopic activation; the dynamic spatiotemporal pattern resulting from sensory activation, including the somatosensory, olfactory, auditory, and visual modalities; and motor preparation and the properties of spontaneously occurring population activity. In this introduction, we focus on VSDI in vivo and review results obtained mostly in the visual system in our laboratory.

A BOLD Assumption

Interpreting fMRI data relies on the assumption that hemodynamic responses reflect neuronal activity. Some recently reported results seem to suggest that this assumption might be less robust than what has been thought so far. Data by Schummers et al. (2008) suggest that hemodynamic responses depend on functional properties of astrocytes as mediators of neuronal activity to blood vessels, and therefore reflect neuronal tuning properties only indirectly. The question is how much the final outcome differs from a linear integration of the local neuronal responses.