# 1 Introduction

## 1 Introduction

Throughout the nervous system, information is encoded in sequences of identical action potentials or spikes. The representation of sense data by these spike trains has been studied for seventy years , but there remain many open questions about the structure of this code. A full understanding of the code requires that we identify its elementary symbols and that we characterize the messages which these symbols represent. Many different possible elementary symbols have been considered, implicitly or explicitly, in previous work on neural coding. These might be the numbers of spikes in time windows of fixed size, or alternatively the individual spikes themselves might be the building blocks of the code. In cells that produce bursts of action potentials, these bursts might be special symbols that convey information in addition to that carried by single spikes. Yet another possibility is that patterns of spikes—across time in one cell or across a population of cells—can have a special significance; this last possibility has received renewed attention as techniques emerge for recording the activity of many neurons simultaneously.

In many methods of analysis, questions about the symbolic structure of the code are mixed with questions about what the symbols represent. Thus, in trying to characterize the feature selectivity of neurons, one often makes the a priori choice to measure the neural response as the spike count or rate in a fixed window. Conversely, in trying to assess the significance of synchronous spikes from pairs of neurons, or bursts of spikes from a single neuron, one might search for a correlation between these events and some particular stimulus features. In each case conclusions about one aspect of the code are limited by assumptions about another aspect. Here we show that questions about the symbolic structure of the neural code can be separated out and answered in an information theoretic framework, using data from suitably designed experiments. This framework allows us to address directly the significance of spike patterns or other compound spiking events: How much information is carried by a compound event? Is there redundancy or synergy among the individual spikes? Are particular patterns of spikes especially informative?

Methods to assess the significance of spike patterns in the neural code share a common intuitive basis:

* Patterns of spikes can play a role in representing stimuli if and only if the occurrence of patterns is linked to stimulus variations.
* The patterns have a special role only if this correlation between sensory signals and patterns is not decomposable into separate correlations between the signals and the pieces of the pattern, e.g. the individual spikes.

We believe that these statements are not controversial. Difficulties arise when we try to quantify this intuitive picture: What is the correct measure of correlation? How much correlation is significant? Can we make statements independent of models and elaborate null hypotheses?

The central claim of this paper is that many of these difficulties can be resolved by careful use of ideas from information theory. Shannon  proved that entropy and information provide the only measures of variability and correlation that are consistent with simple and plausible requirements. Further, while it may be unclear how to interpret, for example, a 20% increase in correlation between spike trains, an extra bit of information carried by patterns of spikes means precisely that these patterns provide a factor of two increase in the ability of the system to distinguish among different sensory inputs. In this work we show see that there is a direct method of measuring the information (in bits) carried by particular patterns of spikes, independent of models for the stimulus features that these patterns might represent. In particular, we can compare the information conveyed by spikes patterns with the information conveyed by the individual spikes that make up the pattern, and determine quantitatively whether the whole is more or less than the sum of its parts.

While this method allows us to compute unambiguously how much information is conveyed by particular patterns, it does not tell us what this information is. Making the distinction between two questions, the structure of the code and the algorithm for translation, we only answer the first of these two. We believe that finding the structural properties of a neural code independent of the translation algorithm is an essential first step towards understanding anything beyond the single spike approximation. The need for identifying the elementary symbols is especially clear when complex multi neuron codes are considered. Moreover, a quantitative measure of the information carried by compound symbols will be useful in the next stage of modeling the encoding algorithm, as a control for the validity of models.

## 2 Formalism

In the framework of information theory  signals are generated by a source with a fixed probability distribution, and encoded into messages by a channel. The coding is probabilistic, and the joint distribution of signals and coded messages determines all quantities of interest; in particular the information transmitted by the channel about the source is an average over this joint distribution. In studying a sensory system, the signals generated by the source are the stimuli presented to the animal, and the messages in the communication channel are sequences of spikes in a neuron or in a population of neurons. Both the stimuli and the spike trains are random variables, and they convey information mutually because they are correlated. The problem of quantifying this information has been discussed from several points of view . Here we address the question of how much information is carried by particular “events” or combinations of action potentials.

### 2.1 Defining information from one event

A discrete event $`E`$ in the spike train is defined as a specific combination of spikes. Examples are a single spike, a pair of spikes separated by a given time, spikes from two neurons that occur in synchrony, and so on. Information is carried by the occurrence of events at particular times and not others, implying that they are correlated with some stimulus features and not with others. Our task is to express this information in terms of quantities that are easily measured experimentally.

In experiments, as in nature, the animal is exposed to stimuli at each instant of time. We can describe this sensory input by a function $`s(t^{})`$, which may have many components to parameterize the time dependence of multiple stimulus features. In general, the information gained about $`s(t^{})`$ by observing a set of neural responses is

$`I`$ $`=`$ $`{\displaystyle \underset{\mathrm{responses}}{}}{\displaystyle Ds(t^{})P[s(t^{})\&\mathrm{response}]\mathrm{log}_2\left(\frac{P[s(t^{})\&\mathrm{response}]}{P[s(t^{})]P[\mathrm{response}]}\right)},`$   
where information is measured in bits. It is useful to note that this mutual information can be rewritten in two complementary forms:

$`I`$ $`=`$ $`{\displaystyle Ds(t^{})P[s(t^{})]\mathrm{log}_2P[s(t^{})]}`$ (2)   
$`+{\displaystyle \underset{\mathrm{responses}}{}}P[\mathrm{response}]{\displaystyle Ds(t^{})P[s(t^{})|\mathrm{response}]\mathrm{log}_2P[s(t^{})|\mathrm{response}]}`$   
$`=`$ $`S[P(s)]S[P(s|\mathrm{response})]_{\mathrm{response}},`$   
or

$`I`$ $`=`$ $`{\displaystyle \underset{\mathrm{responses}}{}}P(\mathrm{response})\mathrm{log}_2P(\mathrm{response})`$ (3)   
$`+{\displaystyle Ds(t^{})P[s(t^{})]\underset{\mathrm{responses}}{}P[\mathrm{response}|s(t^{})]\mathrm{log}_2P[\mathrm{response}|s(t^{})]}`$   
$`=`$ $`S[P(\mathrm{response})]S[P(\mathrm{response}|s)]_s,`$   
where $`S`$ denotes the entropy of a distribution; by $`\mathrm{}_s`$ we mean an average over all possible values of the sensory stimulus, weighted by their probability of occurrence, and similarly for $`\mathrm{}_{\mathrm{responses}}`$. In the first form, Eq. (2), we focus on what the responses are telling us about the sensory stimulus : different responses point more or less reliably to different signals, and our average uncertainty about the sensory signal is reduced by observing the neural response. In the second form, Eq. (3), we focus on the variability and reproducibility of the neural response . The range of possible responses provides the system with a capacity to transmit information, and the variability which remains when the sensory stimulus is specified constitutes noise; the difference between the capacity and the noise is the information.

We would like to apply this second form to the case where the neural response is a particular type of event. When we observe an event $`E`$, information is carried by the fact that it occurs at some particular time $`t_E`$. The range of possible responses is then the range of times $`0<t_E<T`$ in our observation window. Alternatively, when we observe the response in a particular small time bin of size $`\mathrm{\Delta }t`$, information is carried by the fact that the event $`E`$ either occurs or does not. The range of possible responses then includes just two possibilities. Both of these points of view have an arbitrary element: the choice of bin size $`\mathrm{\Delta }t`$ and the window size $`T`$. Characterizing the properties of the system, as opposed to our observation power, requires taking the limit of high time resolution ($`\mathrm{\Delta }t0`$) and long observation times ($`T\mathrm{}`$). As will be shown in the next section, in this limit the two points of view give the same answer for the information carried by an event.

### 2.2 Information and event rates

A crucial role is played by the event rate $`r_E(t)`$, the probability per unit time that an event of type $`E`$ occurs at time $`t`$, given the stimulus history $`s(t^{})`$. Empirical construction of the event rate $`r_E(t)`$ requires repetition of same stimulus history many times, so that a histogram can be formed (see Figure 1). For the case where events are single spikes, this is the familiar time dependent firing rate or post-stimulus time histogram (Figure 1c); the generalization to other types of events is illustrated by Figs. 1c and 1d. Intuitively, a uniform event rate implies that no information is transmitted, whereas the presence of sharply defined features in the event rate implies that much information is transmitted by these events; see, for example, the discussion by Vaadia et al. . We now formalize this intuition, and show how the average information carried by a single event is related quantitatively to the time dependent event rate.

Let us take the first point of view about the neural response variable, in which the range of responses is described by the possible arrival times $`t`$ of the event. What is the probability of finding the event at a particular time $`t`$? Before we know the stimulus history $`s(t^{})`$, all we can say is that the event can occur anywhere in our experimental window of size $`T`$, so that the probability is uniform $`P(\mathrm{response})=P(t)=1/T`$, with an entropy of $`S[P(t)]=\mathrm{log}_2T`$. Once we know the stimulus, we also know the event rate $`r_E(t)`$, and so our uncertainty about the occurrence time of the event is reduced. Events will occur preferentially at times where the event rate is large, so the probability distribution should be proportional to $`r_E(t)`$; with proper normalization $`P(\mathrm{response}|s)=P(t|s)=r_E(t)/(T\overline{r}_E)`$. Then the conditional entropy is

$`S[P(t|s)]`$ $`=`$ $`{\displaystyle _0^T}𝑑tP(t|s)\mathrm{log}_2P(t|s)`$ (4)   
$`=`$ $`{\displaystyle \frac{1}{T}}{\displaystyle _0^T}𝑑t{\displaystyle \frac{r_E(t)}{\overline{r}_E}}\mathrm{log}_2\left({\displaystyle \frac{r_E(t)}{\overline{r}_ET}}\right).`$   
In principle one should average this quantity over different stimuli $`s(t^{})`$, however if the time $`T`$ is long enough and the stimulus history sufficiently rich, it is self-averaging. The reduction in entropy is then the gain in information, so

$`I(E;s)`$ $`=`$ $`S[P(t)]S[P(t|s)]`$ (5)   
$`=`$ $`{\displaystyle \frac{1}{T}}{\displaystyle _0^T}𝑑t\left({\displaystyle \frac{r_E(t)}{\overline{r}_E}}\right)\mathrm{log}_2\left({\displaystyle \frac{r_E(t)}{\overline{r}_E}}\right).`$   
This formula expresses the information conveyed by an event of type $`E`$ as an integral over time, of a quantity which depends only on the responses. There is no explicit dependence on the joint distribution of stimuli and responses; it is implicit that by integrating over time we are in fact sampling the distribution of stimuli, whereas by estimating the function $`r_E(t)`$ as a histogram we are sampling the distribution of the responses given a stimulus. We may write the information equivalently as an average over the stimulus instead of over time,

$`I(E;s)`$ $`=`$ $`\left({\displaystyle \frac{r_E(t)}{\overline{r}_E}}\right)\mathrm{log}_2\left({\displaystyle \frac{r_E(t)}{\overline{r}_E}}\right)_s,`$ (6)   
where here the average is over all possible value of $`s`$ weighted by their probabilities $`P(s)`$.

In the second view, the neural response is a binary random variable, $`\sigma _E\{0,1\}`$, marking the occurrence or non occurrence of an event of type $`E`$ in a small time bin of size $`\mathrm{\Delta }t`$. Suppose, for simplicity, that the stimulus takes on a finite set of values $`s`$ with probabilities $`P(s)`$. These in turn induce the event $`E`$ with probability $`p_E(s)=P(\sigma _E=1|s)=r_E(s)\mathrm{\Delta }t`$, with an average probability for the occurrence of the event $`\overline{p}_E=_sP(s)r_E(s)\mathrm{\Delta }t=\overline{r}_E\mathrm{\Delta }t`$. The information is the difference between the prior entropy and the conditional entropy: $`I(E;s)=S(s)S(s|\sigma _E)`$, where the conditional entropy is an average over the two possible values of $`\sigma _E`$. The conditional probabilities are found from Bayes’ rule,

$`\text{P}(s|\sigma _E=1)`$ $`=`$ $`{\displaystyle \frac{P(s)p_E(s)}{\overline{p}_E}}`$   
$`\text{P}(s|\sigma _E=0)`$ $`=`$ $`{\displaystyle \frac{P(s)(1p_E(s))}{(1\overline{p}_E)}},`$ (7)   
and with these one finds the information, 

$`I(E;s)`$ $`=`$ $`{\displaystyle \underset{s}{}}P(s)\mathrm{log}_2P(s)+{\displaystyle \underset{\sigma _E=0,1}{}}P(s|\sigma _E)\mathrm{log}_sP(s|\sigma _E)`$ (8)   
$`=`$ $`{\displaystyle \underset{s}{}}P(s)\left[p_E(s)\mathrm{log}_2\left({\displaystyle \frac{p_E(s)}{\overline{p}_E}}\right)+(1p_E(s))\mathrm{log}_2\left({\displaystyle \frac{1p_E(s)}{1\overline{p_E}}}\right)\right].`$   
This expression is again an average over all stimulus values, of a property which only depends on the responses. Taking the limit $`\mathrm{\Delta }t0`$, consistent with the requirement that the event can occur at most once, one finds the average information conveyed in a small time bin; dividing by the average probability of an event one obtains Eq. (6) as the information per event.

Equation (6), and its time averaged form (5), is an exact formula which can be used in any situation where a rich stimulus history can be presented repeatedly. It enables the evaluation of the information for arbitrarily complex events, independent of assumptions about the encoding algorithm.

Let us consider in more detail the simple case where events are single spikes. Then the average information conveyed by a single spike becomes an integral over the time dependent spike rate $`r(t)`$,

$$I(1\mathrm{spike};\mathrm{s})=\frac{1}{T}_0^T𝑑t\left(\frac{r(t)}{\overline{r}}\right)\mathrm{log}_2\left(\frac{r(t)}{\overline{r}}\right).$$
(9)   
It makes sense that the information carried by single spikes should be related to the spike rate, since this rate as a function of time gives a complete description of the ‘one body’ statistics of the spike train, in the same way that the single particle density describes the one body statistics of a gas or liquid. Several previous works have noted this relation, and the formula (9) has an interesting history. If the spike train is a modulated Poisson process, then Eq. (9) provides an upper bound on information transmission (per spike) by the spike train as a whole . In studying the coding of location by cells in the rat hippocampus, Skaggs et al.  assumed that successive spikes carried independent information, and that the spike rate was determined by the instantaneous location, and obtained Eq. (9) with the time average replaced by an average over locations. DeWeese  showed that the rate of information transmission by a spike train could be expanded in a series of integrals over correlation functions, where successive terms would be small if the number of spikes per correlation time were small; the leading term, which would be exact if spikes were uncorrelated, is Eq. (9). Panzeri et al.  show that Eq. (9), multiplied by the mean spike rate to give an information rate (bits/s), is the correct information rate if we count spikes in very brief segments of the neural response, which is equivalent to asking for the information carried by single spikes. For further discussion of the relation to previous work, see Appendix A.

A crucial point here is the generalization to Eq. (5), and this result applies to the information content of any point events in the neural response—pairs of spikes with a certain separation, coincident spikes from two cells, … —not just single spikes. Moreover, in the analysis of experiments we will emphasize the use of this formula as an exact result for the information content of single events, rather than an approximate result for the spike train as a whole, and this approach will enable us to address questions concerning the structure of the code and the role played by various point events.

## 3 Experiments in the fly visual system

In this section, we use our formalism to analyze experiments on the movement sensitive cell H1 in the visual system of the blowfly Calliphora vicina. We address the issue of the information conveyed by pairs of spikes in this neuron, as compared to the information conveyed independently by single spikes. The quantitative results of this section —numbers for the information, effects of synergy and redundancy among spikes— are specific to this system and to the stimulus conditions used. The theoretical approach, however, is valid generally and can be applied similarly to other experimental systems, to find out the significance of various patterns in single cells or in a population.

### 3.1 Synergy between spikes

The experimental setup described in Appendix B gives us control over the input and output of the H1 neuron in the fly. The horizontal motion across the visual field is the input sensory stimulus $`s(t)`$, which we draw from a probability distribution $`P(s)`$, and the spike train recorded from H1 is the neural response. Figure 1a shows a segment of the stimulus presented to the fly, and 1b illustrates the response to many repeated presentations of this segment. The histogram of spike times across the ensemble of repetitions provides an estimate of the spike rate $`r(t)`$ (Fig. 1c), and Eq. (5) gives the information carried by a single spike, $`I(1\mathrm{spike};s)=1.53\pm 0.05\mathrm{bits}`$. Figure 2 illustrates the details of how the formula was used, with an emphasis on the effects of finiteness of the data. In this experiment, a stimulus of length $`T=10`$ sec was repeated 350 times. As seen from Figure 2, a stable result could be obtained from a smaller number of repetitions.

If each spike were to convey information independently, then with the mean spike rate $`\overline{r}=37`$ spikes/s, the total information rate would be $`R_{\mathrm{info}}=56`$ bits/s. We used the variability and reproducibility of continuous segments in the neural response , in order to estimate the total information rate in the spike train in this experiment, and found that $`R_{\mathrm{info}}=75`$ bits/s. Thus, the information conveyed by the spike train as a whole is larger than the sum of contributions from individual spikes, indicating cooperative information transmission by patterns of spikes in time. This synergy among spikes motivates the search for especially informative patterns in the spike train.

We consider compound events that consist of two spikes separated by a time $`\tau `$, with no constraints on what happens between them. Figure 1 shows segments of the event rate $`r_\tau (t)`$ for $`\tau =3(\pm 1)`$ ms (Fig. 1d), and for $`\tau =17(\pm 1)`$ ms (Fig. 1e). The information carried by spike pairs as a function of the interspike time $`\tau `$, computed from Eq. (5), is shown in Fig. 3. For large $`\tau `$ spikes contribute independent information, as expected. This independence is established within $`3040`$ ms, comparable to the behavioral response times of the fly . There is a mild redundancy ($`1020\%`$) at intermediate separations, and a very large synergy (up to $`130\%`$) at small $`\tau `$.

Related results were obtained using the correlation of spike patterns with stimulus features . There the information carried by spike patterns was estimated from the distribution of stimuli given each pattern, thus constructing a statistical model of what the patterns “stand for” (see details in Appendix A). Since the time dependent stimulus is in general of high dimensionality, its distribution cannot be sampled directly and some approximations must be made. de Ruyter van Steveninck and Bialek  made the approximation that patterns of a few spikes encode projections of the stimulus onto low dimensional subspaces, and the information carried by such patterns was evaluated only in this subspace. The informations obtained in this approximation are bounded from above by the true information carried by the patterns, as estimated directly with the methods presented here.

### 3.2 Origins of synergy

Synergy means, quite literally, that two spikes together tell us more than two spikes separately. Synergistic coding is often discussed for populations of cells, where extra information is conveyed by patterns of coincident spikes from several neurons , while here we see direct evidence for extra information in pairs of spikes across time. The mathematical framework for describing these effects is the same, and a natural question is: what are the conditions for synergistic coding?

The average synergy $`\mathrm{Syn}[E_1,E_2;s]`$ between two events $`E_1`$ and $`E_2`$ is the difference between the information about the stimulus $`s`$ conveyed by the pair, and the information conveyed by the two events independently,

$$\mathrm{Syn}[E_1,E_2;s]=I[E_1,E_2;s](I[E_1;s]+I[E_2;s]).$$
(10)   
We can rewrite the synergy as:

$$\mathrm{Syn}[E_1,E_2;s]=I[E_1;E_2|s]I[E_1;E_2].$$
(11)   
The first term is the mutual information between the events computed across an ensemble of repeated presentations of the same stimulus history. It describes the gain in information due to the locking of compound event $`(E_1,E_2)`$ to particular stimulus features. If events $`E_1`$ and $`E_2`$ are correlated individually with the stimulus but not with one another, this term will be zero, and these events cannot be synergistic on average. The second term is the mutual information between events when the stimulus is not constrained, or equivalently the predictability of event $`E_2`$ from $`E_1`$. This predictability limits the capacity of $`E_2`$ to carry information beyond that already conveyed by $`E_1`$. Synergistic coding ($`\mathrm{Syn}>0`$) thus requires that the mutual information among the spikes is increased by specifying the stimulus, which makes precise the intuitive idea of ‘stimulus dependent correlations’.

Returning to our experimental example, we identify the events $`E_1`$ and $`E_2`$ as the arrivals of two spikes, and consider the synergy between them as a function of the time $`\tau `$ between them. In terms of event rates, we compute the information carried by a pair of spikes separated by a time $`\tau `$, Eq. (5), as well as the information carried by two individual spikes. The difference between these two quantities is the synergy between two spikes, which can be written as

$`\mathrm{Syn}(\tau )`$ $`=`$ $`\mathrm{log}_2\left({\displaystyle \frac{\overline{r}_\tau }{\overline{r}^2}}\right)+{\displaystyle \frac{1}{T}}{\displaystyle _0^T}𝑑t{\displaystyle \frac{r_\tau (t)}{\overline{r}_\tau }}\mathrm{log}_2\left[{\displaystyle \frac{r_\tau (t)}{r(t)r(t\tau )}}\right]`$ (12)   
$`+{\displaystyle \frac{1}{T}}{\displaystyle _0^T}𝑑t\left[{\displaystyle \frac{r_\tau (t)}{\overline{r}_\tau }}+{\displaystyle \frac{r_\tau (t+\tau )}{\overline{r}_\tau }}2{\displaystyle \frac{r(t)}{\overline{r}}}\right]\mathrm{log}_2[r(t)].`$   
The first term in this equation is the logarithm of the normalized correlation function, and hence measures the rarity of spike pairs with separation $`\tau `$; the average of this term over $`\tau `$ is the mutual information between events in Eq. (11). The second term is related to the local correlation function and measures the extent to which the stimulus modulates the likelihood of spike pairs. The average of this term over $`\tau `$ gives the mutual information conditional on knowledge of the stimulus \[the first term in Eq. (11)\]. The average of the third term over $`\tau `$ is zero, and numerical evaluation of this term from the data shows that it is negligible at most values of $`\tau `$.

We thus find that the synergy between spikes is approximately a sum of two terms, whose averages over $`\tau `$ are the terms in Eq. (11). A spike pair with a separation $`\tau `$ then has two types of contributions to the extra information it carries: the two spikes can be correlated conditional on the stimulus, or the pair could be a rare and thus surprising event. The rarity of brief pairs is related to neural refractoriness, but this effect alone is insufficient to enhance information transmission; the rare events must also be related reliably to the stimulus. In fact, conditional on the stimulus, the spikes in rare pairs are strongly correlated with each other, and this is visible in Fig. 1a: from trial to trial, adjacent spikes jitter together as if connected by a stiff spring. To quantify this effect, we find for each spike in one trial the closest spike in successive trials, and measure the variance of the arrival times of these spikes. Similarly, we measure the variance of the interspike times. Figure 4a shows the ratio of the interspike time variance to the sum of the arrival time variances of the spikes that make up the pair. For large separations this ratio is unity, as expected if spikes are locked independently to the stimulus, but as the two spikes come closer it falls below one quarter.

Both the conditional correlation among the members of the pair (Fig. 4a) and the relative synergy (Fig. 4b) depend strongly on the interspike separation. This dependence is nearly invariant to changes in image contrast, although the spike rate and other statistical properties are strongly affected by such changes. Brief spike pairs seem to retain their identity as specially informative symbols over a range of input ensembles. If particular temporal patterns are especially informative, then we would lose information if we failed to distinguish among different patterns. Thus there are two notions of time resolution for spike pairs: the time resolution with which the interspike time is defined, and the absolute time resolution with which the event is marked. Figure 5 shows that, for small interspike times, the information is much more sensitive to changes in the interspike time resolution (open symbols) than to the absolute time resolution (filled symbols). This is related to the slope in Figure 2: in regions where the slope is large, events should be finely distinguished in order to retain the information.

### 3.3 Implications of synergy

The importance of spike timing in the neural code has been under debate for some time now. We believe that some issues in this debate can be clarified using a direct information theoretic approach. Following MacKay and McCulloch , we know that marking spike arrival times with higher resolution provides an increased capacity for information transmission. The work of Strong et al.  shows that for the fly’s H1 neuron, the increased capacity associated with spike timing is indeed used with nearly constant efficiency down to millisecond resolution. This efficiency can be the result of a tight locking of individual spikes to a rapidly varying stimulus, and it could also be the result of temporal patterns providing information beyond rapid rate modulations. The analysis given here shows that for H1, pairs of spikes can provide much more information than two individual spikes, information transmission is much more sensitive to the relative timing of spikes than to their absolute timing, and these synergistic effects survive averaging over all similar patterns in an experiment. On the time scales of relevance to fly behavior, the amount of synergy among spikes in H1 allows this single cell to provide an extra factor of two in resolving power for distinguishing different trajectories of motion across the visual field.

## 4 Summary

In summary, information theory allows us to quantify the symbolic structure of a neural code independent of the rules for translating between spikes and stimuli. In particular, this approach tests directly the idea that patterns of spikes are special events in the code, carrying more information than expected by adding the contributions from individual spikes. These quantities can be measured directly from data. It is of practical importance that the formulas rely on low order statistical measures of the neural response, and hence do not require enormous data sets to reach meaningful conclusions. The method is of general validity and is applicable to patterns of spikes across a population of neurons, as well as across time.

In our experiments on the fly visual system, we found that an event composed of a pair of spikes can carry far more than the information carried independently by its parts. Two spikes that occur in rapid succession appear to be special symbols that have an integrity beyond the locking of individual spikes to the stimulus. This is analogous to the encoding of sounds in written English: the symbols ‘th,’ ‘sh,’ and ‘ch’ are each elementary and stand for sounds that are not decomposable into sounds represented by each of the constituent letters. For such pairs to act effectively as special symbols, mechanisms for ‘reading’ them must exist at subsequent levels of processing. Synaptic transmission is sensitive to interspike times in the 2 – 20 ms range , and it is natural to suggest that synaptic mechanisms on this time scale play a role in such reading. Recent work on the mammalian visual system  provides direct evidence that pairs of spikes close together in time can be especially efficient in driving postsynaptic neurons.

## Acknowledgements

We thank G. Lewen and A. Schweitzer for their help with the experiments and N. Tishby for many helpful discussions. Work at the IAS was supported in part by DOE grant DE–FG02–90ER40542, and work at the IFSC was supported by the Brazilian agencies FAPESP and CNPq.   
Appendix A: Relation to previous work   
Patterns of spikes and their relation to sensory stimuli have been quantified in the past through the use of correlation functions. The event rates that we have defined here, which are directly connected to the information carried by patterns of spikes by Eq. (5), are in fact just properly normalized correlation functions. The event rate for pairs of spikes from two separate neurons is related to the joint post-stimulus time histogram defined by Aertsen and coworkers  Making this connection explicit is also an opportunity to see how the present formalism applies to events defined across two cells.

Consider two cells, $`A`$ and $`B`$, generating spikes at times $`\{t_\mathrm{i}^A\}`$ and $`\{t_\mathrm{i}^B\}`$, respectively. It will be useful to think of the spike trains as sums of unit impulses at the spike times,

$`\rho ^A(t)`$ $`=`$ $`{\displaystyle \underset{\mathrm{i}}{}}\delta (tt_\mathrm{i}^A)`$ (A.1)   
$`\rho ^B(t)`$ $`=`$ $`{\displaystyle \underset{\mathrm{i}}{}}\delta (tt_\mathrm{i}^B).`$ (A.2)   
Then the time dependent spike rates for the two cells are

$`r^A(t)`$ $`=`$ $`\rho ^A(t)_{\mathrm{trials}},`$ (A.3)   
$`r^B(t)`$ $`=`$ $`\rho ^B(t)_{\mathrm{trials}},`$ (A.4)   
where $`\mathrm{}_{\mathrm{trials}}`$ denotes an average over multiple trials in which the same time dependent stimulus $`s(t^{})`$ is presented. These spike rates are the probabilities per unit time for the occurrence of a single spike in either cell $`A`$ or cell $`B`$, also called the post-stimulus time histogram (PSTH). We can define the probability per unit time for a spike in cell $`A`$ to occur at time $`t`$ and a spike in cell $`B`$ to occur at time $`t^{}`$, and this will be the joint post-stimulus time histogram,

$$\mathrm{JPSTH}^{AB}(t,t^{})=\rho ^A(t)\rho ^B(t^{})_{\mathrm{trials}}.$$
(A.5)   
Alternatively, we can consider an event $`E`$ defined by a spike in cell $`A`$ at time $`t`$ and a spike in cell $`B`$ at time $`t\tau `$, with the relative time $`\tau `$ measured to a precision of $`\mathrm{\Delta }\tau `$. Then the rate of these events is

$`r_E(t)`$ $`=`$ $`{\displaystyle _{\mathrm{\Delta }\tau /2}^{\mathrm{\Delta }\tau /2}}𝑑t^{}\mathrm{JPSTH}^{AB}(t,t\tau +t^{})`$ (A.6)   
$``$ $`\mathrm{\Delta }\tau \mathrm{JPSTH}^{AB}(t,t\tau ),`$ (A.7)   
where the last approximation is valid if our time resolution is sufficiently high. Applying our general formula for the information carried by single events, Eq. (5), the information carried by pairs of spikes from two cells can be written as an integral over diagonal “strips” of the $`\mathrm{JPSTH}`$ matrix,

$$I(E;s)=\frac{1}{T}_0^T𝑑t\frac{\mathrm{JPSTH}^{AB}(t,t\tau )}{\mathrm{JPSTH}^{AB}(t,t\tau )_t}\mathrm{log}_2\left[\frac{\mathrm{JPSTH}^{AB}(t,t\tau )}{\mathrm{JPSTH}^{AB}(t,t\tau )_t}\right],$$
(A.8)   
where $`\mathrm{JPSTH}^{AB}(t,t\tau )_t`$ is an average of the $`\mathrm{JPSTH}`$ over time, which is equivalent to the standard correlation function between the two spike trains.

The discussion by Vaadia et al.  emphasizes that modulations of the $`\mathrm{JPSTH}`$ along the diagonal strips allows correlated firing events to convey information about sensory signals or behavioral states, and this information is quantified by Eq. (A.8). The information carried by the individual cells is related to the corresponding integrals over spike rates, Eq. (9). The difference between the the information conveyed by the compound spiking events $`E`$, and the informations conveyed by spikes in the two cells independently, is precisely the synergy between the two cells at the given time lag $`\tau `$. For $`\tau =0`$, it is the synergy –or extra information– conveyed by synchronous firing of the two cells.

We would like to connect the present approach also with previous work which focused on how events reduce our uncertainty about the stimulus . Before we observe the neural response, all we know is that stimuli are chosen from a distribution $`P[s(t^{})]`$. When we observe an event $`E`$ at time $`t_E`$, this should tell us something about the stimulus in the neighborhood of this time, and this knowledge is described by the conditional distribution $`P[s(t^{})|t_E]`$. If we go back to the definition of the mutual information between responses and stimuli, we can write the information conveyed by one event in terms this conditional distribution,

$`I(E;s)`$ $`=`$ $`{\displaystyle Ds(t^{})𝑑t_EP[s(t^{}),t_E]\mathrm{log}_2\left(\frac{P[s(t^{}),t_E]}{P[s(t^{})]P[t_E]}\right)}`$ (A.9)   
$`=`$ $`{\displaystyle 𝑑t_EP[t_E]Ds(t^{})P[s(t^{})|t_E]\mathrm{log}_2\left(\frac{P[s(t^{})|t_E]}{P[s(t^{})]}\right)}.`$   
If the system is stationary then the coding should be invariant under time translations:

$$P[s(t^{})|t_E]=P[s(t^{}+\mathrm{\Delta }t^{})|t_E+\mathrm{\Delta }t^{}].$$
(A.10)   
This invariance means that the integral over stimuli in Eq. (A.9) is independent of the event arrival time $`t_E`$, so we can simplify our expression for the information carried by a single event,

$`I(E;s)`$ $`=`$ $`{\displaystyle 𝑑t_EP[t_E]Ds(t^{})P[s(t^{})|t_E]\mathrm{log}_2\left(\frac{P[s(t^{})|t_E]}{P[s(t^{})]}\right)}`$ (A.11)   
$`=`$ $`{\displaystyle Ds(t^{})P[s(t^{})|t_E]\mathrm{log}_2\left(\frac{P[s(t^{})|t_E]}{P[s(t^{})]}\right)}.`$   
This formula was used by de Ruyter van Steveninck and Bialek  To connect with the present work, we express the information in Eq. (A.11) as an average over the stimulus,

$`I(E;s)=\left({\displaystyle \frac{P[s(t^{})|t_E]}{P[s(t^{})]}}\right)\mathrm{log}_2\left({\displaystyle \frac{P[s(t^{})|t_E]}{P[s(t^{})]}}\right)_s.`$ (A.12)   
Using Bayes’ rule,

$$\frac{P[s(t^{})|t_E]}{P[s(t^{})]}=\frac{P[t_E|s(t^{})]}{P[t_E]}=\frac{r_E(t_E)}{\overline{r}_E}$$
(A.13)   
where the last term is a result of the distributions of event arrival times being proportional to the event rates, as defined above. Substituting the back to Eq. (A.11), one finds the equivalent of Eq. (6).   
Appendix B: Experimental setup   
In the experiment we used a female blowfly, which was a first generation offspring of a wild fly caught outside. The fly was put inside a plastic tube and immobilized with wax, with the head protruding out. The proboscis was left free so that the fly could be fed regularly with some sugar water. A small hole was cut in the back of the head, close to the midline on the right side. Through this hole, a tungsten electrode was advanced into the lobula plate. This area, which is several layers back from the compound eye, includes a group of large motion detector neurons with wide receptive fields and strong direction selectivity. We recorded spikes extracellularly from one of these, the contralateral H1 neuron . The electrode was positioned such that spikes from H1 could be discriminated reliably, and converted into TTL pulses by a simple threshold discriminator. The TTL pulses fed into a CED 1401 interface, which time stamped the digitized spikes at 10 $`\mu s`$ resolution. To keep exact synchrony over the duration of the experiment, the spike timing clock was derived from the same internal CED 1401 clock that defined the frame times of the visual stimulus.

The stimulus was an rigidly moving bar pattern, displayed on a Tektronix 608 high brightness display. The radiance at average intensity was about 20 mW/(m<sup>2</sup> $``$ sr), which amounts to about 5 $`10^4`$ effectively transduced photons per photoreceptor per second . The bars were oriented vertically, with intensities chosen at random to be $`\overline{I}(1\pm C)`$, where $`C`$ is the contrast. The distance between the fly and the screen was adjusted so that angular subtense of a bar equaled the horizontal interommatidial angle in the stimulated part of the compound eye. This setting was found by determining the eye’s spatial Nyquist frequency through the reverse reaction . For this fly, the horizontal interommatidial angle was 1.45, and the distance to the screen 105 mm. The fly viewed the display through a round 80 mm diameter diaphragm, showing approximately 30 bars. From this we estimate the number of stimulated ommatidia in the eye’s hexagonal raster to be about 612.

Frames of the stimulus pattern were refreshed every 2 ms, and with each new frame the pattern was displayed at a new position. This resulted in an apparent horizontal motion of the bar pattern, which is suitable to excite the H1 neuron. The pattern position was defined by a pseudorandom sequence, simulating independent random numbers uniformly distributed between -0.47 to + 0.47 (equivalent to -0.32 to +0.32 omm, horizontal ommatidial spacings). This corresponds to a diffusion constant of 18.1()<sup>2</sup>/s or 8.6 omm<sup>2</sup>/s. The sequence of pseudorandom numbers contained a repeating part and a nonrepeating part, each 10 seconds long, with the same statistical parameters. Thus in each 20 second cycle the fly saw a 10 second movie that it had seen 20 seconds before, followed by a 10 second movie that was generated independently.

Figures

Fig. 1. Generalized event rates in the stimulus–conditional response ensemble. A time dependent visual stimulus is shown to the fly (a), with the time axis defined to be zero at the beginning of the stimulus. This stimulus runs for 10 s, and is repeatedly presented 360 times. The responses of the H1 neuron to 60 repetitions are shown as a raster (b), in which each dot represents a single spike. From these responses, time dependent event rates $`r_E(t)`$ are estimated: the firing rate (post-stimulus time histogram) (c); the rate for spike pairs with interspike time $`\tau =3\pm 1`$ ms (d) and for pairs with $`\tau =17\pm 1`$ ms (e). These rates allow us to compute directly the information transmitted by the events, using Eq. (5).   

Fig. 2. Finite size effects in the estimation of the information conveyed by single spikes. (a) Information as a function of the bin size $`\mathrm{\Delta }t`$ used for computing the time dependent rate $`r(t)`$ from all 350 repetitions (circles), and from 100 of the repetitions (filled triangles). A linear extrapolation to the limit $`\mathrm{\Delta }t0`$ is shown for the case where all repetitions were used (solid line). (b) Information as a function of the inverse number of repetitions $`N`$, for a fixed bin size $`\mathrm{\Delta }t=2`$ms. (c) Statistical error due to the finiteness of the time segment of length $`T`$. Information shown as a function of the inverse time segment $`1/T`$, was obtained by dividing the full 10-sec segments into smaller segments of length $`T`$ (circles). For each such division, the errorbars represent the standard deviation of the values obtained from the different time intervals. These error bars should follow a square-root law ($`\sigma 1/\sqrt{T}`$) if the small segments are independent. The dashed line shows the best power law fit to the sequence of standard deviations, which extrapolates to an errorbar of $`\sigma 0.05`$ for the full 10-sec segment.   

Fig. 3. Information about the signal transmitted by pairs of spikes, computed from Eq. (5), as a function of the time separation between the two spikes. The dotted line shows the information that would be transmitted by the two spikes independently (twice the single spike information).   

Fig. 4. (a) Ratio between the variance of interspike time and the sum of variances of the two spike times. Variances are measured across repeated presentations of same stimulus, as explained in the text. This ratio is plotted as a function of the interspike time $`\tau `$, for two experiments with different image contrast. (b) Extra information conveyed cooperatively by pairs of spikes, expressed as a fraction of the information conveyed by the two spikes independently. While the single spike information varies with contrast, (1.5 bits/spike for c=0.1 compared to 1.3 bits/spike for c=1), the fractional synergy is almost contrast independent.   

Fig. 5. Information conveyed by spike pairs as a function of time resolution. An event –pair of spikes– can be described by two times: the separation between spikes (relative time), and the occurrence time of the event with respect to the stimulus (absolute time). The information carried by the pair depends on the time resolution in these two dimensions, both specified by the bin size $`\mathrm{\Delta }t`$. Open symbols are measurements of the information for a fixed absolute-time resolution of 2 ms, and a variable relative-time resolution $`\mathrm{\Delta }t`$. Closed symbols correspond to a fixed relative-time resolution of 2 ms, and a variable absolute-time resolution $`\mathrm{\Delta }t`$. For short intervals, the sensitivity to coarsening of the relative time resolution is much greater than to coarsening of the absolute time resolution In contrast, sensitivity to relative and absolute time resolution is the same for the longer, nonsynergistic, interspike separations.